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Review
Pumps through the Ages
Yunyan Qiu,1 Yuanning Feng,1 Qing-Hui Guo,1 R. Dean Astumian,2,* and J. Fraser Stoddart1,3,4,*

SUMMARY The Bigger Picture

Challenges and opportunities:
Artificial molecular machines (AMMs) have profoundly enhanced sci-
entists’ capability to manipulate the relative movements of compo-
The ability to harness artificial
nents within molecules. Mechanically interlocked molecules (MIMs) molecular pumps to produce
with movable components have contributed to the design and syn- away-from-equilibrium
thesis of AMMs. The operation of these wholly synthetic molecular rotaxanes and polyrotaxanes
machines away from equilibrium is governed by ratchet mechanisms with exquisite control over the
that require a supply of energy. In this review, we discuss the emer- numbers, sequences, and
gence of pumps, both natural and crafted through the ages, focusing functions of the threaded rings
on recent advances toward the design and synthesis of artificial mo- has far-reaching consequences
lecular pumps (AMPs) that are capable of creating local concentra- for molecular nanotechnology.
tions of rings on collecting chains. We conclude our discussion by
Design of molecular pumps
considering a recently reported catalysis-driven AMM and the rami- highlights the importance of
fications for exploiting catalysis to control non-equilibrium behavior. kinetics for non-equilibrium
function. The importance of
INTRODUCTION kinetics is particularly evident in
Human history is encapsulated by making and using better tools. The creation and the pumping-cassette design
operation of tools and machines set us apart, more than anything else, from other borrowed from biology where
living species. The innovation of macroscopic machines to manipulate our environ- modulation of barriers can
ment has fundamentally revolutionized daily life. In addition, the exploration of the maintain a non-equilibrium
microscopic natural world in recent centuries has revealed the staggering complexity steady state.
of biomolecular machines responsible for sustaining life, and discoveries have
Controlled motions associated
mounted at an ever-increasing pace during the past seven decades. These complex with mechanical bonding, in
biomolecular machines have been a source of inspiration for scientists and engineers combination with the
to create their artificial counterparts.1–13 Both physicists and chemists have espoused nanotopology that
a long fascination with designing and crafting—from the bottom-up—wholly syn- characterizes artificial knots,
thetic molecular machines.14 With their relative structural simplicity they offer, in offer unprecedented
combination, unlimited possibilities for achieving functional complexity that may opportunities to access high-
someday rival that found in biomolecular machines. The resulting systems chemis- energy conformations in
try15–18 may well provide insight into the origin of life.16,19,20 The ability to pursue nanoconfined environments
chemistry away from equilibrium and release it from the shackles of thermodynamic with profound consequences
control, through the design and construction of artificial molecular machines13,14,21– for all aspects of chemical
24
(AMMs), is a tempting prospect that has been stimulated by the award of the 2016 reactivity.
Nobel Prize in Chemistry to three pioneers of the field of AMMs, namely Jean-Pierre
Sauvage,25 Ben L. Feringa,26 and one of the authors27 of this review.

Since the first report28 of a molecular shuttle in 1991, significant advances have been
made in the design, synthesis, and operating principles that define artificial molec-
ular switches29 and AMMs.13,14,21 Mechanically interlocked molecules25,27,30 (MIMs),
such as rotaxanes and catenanes, where rotaxanes contain a linear dumbbell on
which one or more rings have been threaded and catenanes consist of at least two
mechanically interlocked rings, have served as springboards for the design and syn-
thesis of artificial molecular switches,29 which in turn have found themselves at the
epicenter of a hugely challenging area of research. Over the past few decades,

1952 Chem 6, 1952–1977, August 6, 2020 ª 2020 Elsevier Inc.

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MIMs have been adopted and employed widely, by many practitioners31 of the field,
as the basis for AMMs on account of the ability of their components to undergo high-
ly controlled relative motions in the presence of different stimuli. An ever-increasing
range of AMMs has been designed and produced, including, but not limited to, mo-
lecular switches,29 brakes,32 gears,33,34 motors,35–37 elevators,38 muscles,39–41
pumps,42 hoppers,43 zippers,44 ribosome analogs,45 assemblers,46 robotic arms,47
and nanocars.48,49 Among them, the creation42 of artificial molecular pumps
(AMPs) has become, during the past decade, one of the major goals in unnatural
product synthesis.50 In engineering terms, pumps can, given sufficient external en-
ergy, transport materials against local concentration gradients, thus maintaining
the system in a steady state away from thermodynamic equilibrium.51 This attribute
is reminiscent of what biological machines achieve with a much greater complexity
resulting from billions of years of evolution. The resulting non-equilibrium steady
state defines52 the very act of living.53

We begin with an overview of macroscopic pumps,54 and those found in living sys-
tems,55,56 to focus ultimately on wholly synthetic ones42,57–61 created in the research
laboratory. Our discussions will be centered on the underlying design62 and oper-
ating principles63 of AMPs, comparing them in terms of energy inputs and ratcheting
mechanisms.63,64 Future opportunities and promising research trajectories offered
by AMPs will be highlighted with regard to their properties and potential for finding
practical applications in polymer and materials science.65

MACROSCOPIC AND BIOLOGICAL PUMPS

Before delving into the design and syntheses of AMPs, a brief introduction to macro-
scopic mechanical pumps invented throughout human history, and biomolecular
pumps that have evolved in the natural world, will be presented. To some extent,
they have served as sources of inspiration for establishing the design principles of
AMPs. The common feature shared by macroscopic and biological pumps is their
ability to transport materials from low to high energy states, given supplies of fuel.
The operating mechanisms governing how they perform their functions are, how-
ever, fundamentally different from one another.

Macroscopic Pumps
The development54 of pumps as water-lifting devices has played a profound role in
human history, since water is such a vital commodity for sustaining all forms of life
and has been central to the development of all civilizations. A pump66 is a device
that raises or moves liquid. Humans have utilized pumps for almost 4,000 years since
the invention of shaduf,54 known as the first device for water lifting, possibly in
ancient Egypt ca 2000 BC. The shaduf (Figure 1Ai), which was made of wood and
hand-operated at the time of its invention, lifts water from a well or a canal into a
receptacle of some kind. The most common form of a shaduf consisted of a sus- 1Department of Chemistry, Northwestern
University, 2145 Sheridan Road, Evanston, IL
pended rod and a bucket tied to the rod by a rope at one end with a balancing
60208, USA
weight at the other end. Similar designs were used widely throughout the ancient 2Department of Physics, University of Maine,
world. 5709 Bennet Hall, Orono, ME 04469, USA
3Institutefor Molecular Design and Synthesis,
In ancient China, another water-lifting pump54 called lùlú (Figure 1Aii) was invented Tianjin University, Tianjin 300072, P.R. China
4School of Chemistry, University of New South
around 1000 BC in the northern part of China and was used to draw and transport
Wales, Sydney, NSW 2052, Australia
water from rivers over long distances. An invention by Philon of Byzantium, which
*Correspondence:
was a paddle-wheel-driven bucket-chain pump54 (Figure 1Aiii), appeared around astumian@maine.edu (R.D.A.),
280–220 BC. This pump was comparatively more complicated and was used to lift stoddart@northwestern.edu (J.F.S.)
water from rivers to higher grounds. Arguably, one of the greatest inventions of all https://doi.org/10.1016/j.chempr.2020.07.009

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Figure 1. Representative Macroscopic and Biological Pumps

(A) Pictorial and graphical representations of some macroscopic pumps including (Ai) the shaduf (reprinted from Yannopoulos et al., 54 copyright 2015
MDPI), (Aii) the lùlú (reprinted from Yannopoulos et al., 54 copyright 2015 MDPI), (Aiii) the paddle-wheel-driven bucket-chain pump (reprinted from
Yannopoulos et al., 54 copyright 2015 MDPI), (Aiv) the Archimedes screw pump, where water is pushed upward by the screw in the closed chamber. Each
rotation of the crank pumps a fixed volume of water. The pumping can be quantified by plotting the distances between two fiduciary marks on the screw
and some fixed points as shown in the diagram. The volume of water transported is proportional to the area enclosed by the ellipse in the insert in the
upper right-hand corner. (Av) the gear pump, (Avi) the piston pump, and (Avii) the steam-engine-driven pump.
(B) Graphical representations of the crystal structures of several of biological pumps, namely (Bi) Na+ ,K+ -ATPase (image from the RCSB PDB (rcsb.org)
of PDB ID 3KDP 67 ), (Bii) F oF 1 -ATP synthase (Adapted from Guo et al., 68 copyright 2019 eLife Sciences Publications), and (Biii) calcium ATPase shown in
two panels by rotation (adapted with permission from Toyoshima et al., 69 copyright 2000 Springer Nature).

time, a mechanical device called the Archimedes screw pump54 (Figure 1Aiv), which
is still in use to this day, was designed around 200 BC. This pump had an embedded
screw created from a wooden shaft with curved propellers formed of willows or
branches that rotated with respect to the shaft. Rotation of the screw in the appro-
priate direction allows trapped water to be lifted from the lower to the higher end
of the screw. Each rotation of the screw pumps a fixed volume of water by a geomet-
ric pumping mechanism.70 Pumps, such as the gear (Figure 1Av) and piston (Fig-
ure 1Avi) pumps, can all be viewed as descendants of the Archimedes screw
pump, since they all operate under a similar mechanism by positive displacement
in which a fixed volume of liquid is pumped for each cycle of operation. Perhaps
one of the most important piston pumps is the one driven by a steam engine (Fig-
ure 1Avii). Introduced by Thomas Newcomen, an English inventor, in the early

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18th century, this pump was used to remove water from coal mines. The well-under-
stood mechanical principles behind the operation of macroscopic machines are, for
the most part, not applicable in the world of (bio)molecular machines, where high
viscosity renders inertia irrelevant and continual buffeting by thermal noise gives
rise to ineluctable Brownian motion.22,71–73

Biological Pumps
Molecular biophysicists explore how billions of years of evolution have allowed bio-
molecular machines to operate in a precise manner despite the fundamentally
random nature of the Brownian motion72 by which their functions are expressed.
One of the most important biological processes is the transport of ions across the
membranes of cells and organelles74 against their electrochemical gradients, a pre-
requisite for many other biological functions. This active transport is accomplished
by coupling the ion pumps to energy sources such as adenosine triphosphate
(ATP) hydrolysis. The development of modern diffraction,75,76 spectroscopy,77 mi-
croscopy,78–82 and super-resolution83,84 techniques has facilitated our understand-
ing of how these motor proteins operate in a selective manner to transport a range of
ions across many different membranes.

There are many biomolecular pumps—transmembrane proteins that contact both the
intra- and extra-cellular environments. These proteins use chemical energy to move
ions or other molecules from low to high electrochemical potentials, i.e., in the direction
opposite to that expected based solely on equilibrium thermodynamics. The Na+,K+-
ATPase,67 which was originally identified by Skou85 in 1957, a discovery for which he
was ultimately awarded a share of the Nobel Prize in Chemistry 40 years later, represents
(Figure 1Bi) a significant class of membrane-bound ion pumps. At first glance, it would
seem that the pumps, shown in Figure 1B, have nothing in common with their macroscopic
counterparts in Figure 1A. Indeed, the word pump was at first too controversial a descrip-
tion of a biological molecule. Skou in his 1957 paper85 simply referred to the influence of
some cations on an ATPase. Nevertheless, subsequent experiments revealed the role of
Na+,K+-ATPase in using energy from ATP hydrolysis for creating and maintaining gradi-
ents of Na+ and K+ ions across cell membranes. This biological pump will be of central
importance to the exposition at hand since it was the first example of a biomolecular ma-
chine where energy, normally supplied by ATP hydrolysis, could be substituted86 by en-
ergy from an oscillating electric field. Before we begin a detailed consideration of how
this energy transduction occurs, let us mention briefly a few other biomolecular pumps.

The FoF1-ATP synthase68,87 (Figure 1Bii) is a protein found in the mitochondria of

cells and is the chief agent responsible for making the ATP that powers our meta-
bolism and keep us alive. The necessary energy is provided by the exergonic trans-
port of protons down gradients formed by electron transport chain proteins using
the energy of catabolism. The hypothesis—and subsequent demonstration—that
proton transport through the Fo portion of the protein is accompanied by rotation
of the F1 part of the protein, resulted in the award of the 1997 Nobel Prize in Chem-
istry to Paul Boyer88 who unraveled the mechanism known as conformational
coupling and John Walker89 who elucidated the X-ray crystal structure of the com-
plex. More recently, Arieh Warshel90 was awarded the Nobel Prize in Chemistry in
2013, along with Michael Levitt91 and Martin Karplus,92 for developing computa-
tional techniques by which biomolecules can be investigated. Warshel, in particular,
has made significant inroads93,94 toward the understanding of FoF1-ATPase. Finally,
the calcium ATPase69 (Figure 1Biii, in the same class, known as E1,E2 ATPases, as the
Na+,K+-ATPase) has been widely studied, especially by Jencks,95 who focused on
how ATP hydrolysis is coupled to drive uphill transport of Ca2+ from low to high

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Figure 2. Transport Systems Unified by the Concept of Geometric Pumping

(A) The schematic diagram for a positive displacement (geometric) pump used to move water from
a low-level to a high-level reservoir. As the plunger is pushed down, the pressure causes valve 1 to
close and valve 2 to open, so that the water in the syringe is expelled into reservoir 2. When the
plunger is pulled up, the pressure closes valve 2 and opens valve 1 so that the syringe fills with water
from reservoir 1. The process is repeated cyclically, transferring water from reservoir 1 to reservoir 2
despite the lower level of water in reservoir 1 than that in reservoir 2.
(B) Calcium ATPase similarly has two configurations: E 1 in which a high-affinity binding site for Ca2+
ions is exposed to the cytosol and E 2 in which a low-affinity binding site is exposed to the lumen.
(C) Both microscopic and macroscopic pumps can be conceptualized in terms of energy diagrams,
where the relative heights of the barriers indicate which of the gates is open, and which is ‘‘closed’’,
and the energy level, i.e., the affinity, indicates the amount of material bound.
(D) Physiologically the E 1 to E 2 transition is linked to phosphorylation by ATP, but this transition can
also be driven externally, e.g., by an applied electric field. If the relative barrier height u adjusts to
the field more rapidly than does the binding energy ε, the two will, in the presence of an AC field,
oscillate out of phase with one another.
(E) (Ei) Schematic illustration of how the energy profile undergoes a cycle from high-affinity open on
the left / high-affinity open on the right / low-affinity open on the right / low-affinity open on
the left / high-affinity open on the left. (Eii) Plots of the equilibrium occupancy of the well, Qeq ðtÞ =

1
1
1 + eεðtÞ=RT versus the left-right splitting probability FðtÞ = 1 + euðtÞ=RT for low frequency
(dashed), intermediate frequency (solid), and high frequency (dashed dot) relative to the

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Figure 2. Continued
~ + Anonad u
u ~2
conformational relaxation time. (Eiii) The ion flux, given by the expression jIL/R jz 2
, is
1+u ~
+ +
plotted (solid curve), along with experimental data for Na (blue triangle) and Rb (red square), an
analog of K + for which there is a convenient radioactive isotope. The frequency u ~ is the
perturbation frequency divided by the individual ion relaxation time (10 3 s for Rb+ and 10 6 s for
Na +) and the fit parameter Anonad accounts for the fact that the frequency of the external field is not
much less than the inverse relaxation time of the ion passage at very high frequency and is taken to
be Anonad = 0:14.

electrochemical potential. There are many other biomolecular pumps, reflecting the
importance to organisms of the ability to move materials from one compartment to
another in a regulated manner, forming large gradients of ions and small molecules.
These include lactose permease96 in Escherichia coli, which transduces the free en-
ergy in electrochemical proton gradients to drive the accumulation of lactose, and
ABC (ATP binding cassette) transporters,97 which represent a family of membrane-
spanning transport proteins that translocate a range of substrates and contribute
to multidrug resistance.98

In Figure 2, we highlight the conceptual connection between macroscopic geomet-

ric pumps (Figure 2A) and molecular pumps (Figure 2B) using the calcium ATPase as
a specific example. There are two well-defined states, known as E1 and E2, with fea-
tures that can be related to those of the macroscopic syringe pump (Figure 2A). The
state E1, which is open to the cytosol and "closed" to the lumen, has a high affinity for
Ca2+ ions. The state E2, which is open to the lumen and "closed" to the cytosol, has a
low affinity for Ca2+ ions. The word "closed" is in quotation marks. In the microscopic
world, the entry and exit rates can be very small but their ratio must always reflect the
equilibrium constant for the association-cum-dissociation process. See Box 1. These
ideas are illustrated in terms of energy diagrams as shown in Figure 2C.

PUMPING CASSETTE
The Archimedes screw, and all positive displacement pumps, carry out their function
by a geometric mechanism,70 where each cycle pumps a fixed volume of water.
Most, if not all, microscopic pumps work by such a geometric mechanism. David
Thouless,105 one of the 2016 Nobel Laureates in Physics, introduced the idea of
adiabatic geometric pumping of electrons in which the Hamiltonian describing an
electronic system is modulated slowly enough that no energy is dissipated and yet
there is a net current. This electron current is quantized and so the effect has found
importance in metrology as a fundamental quantification of the Coulomb. Similar ef-
fects apply at the level of single cells.106 As Purcell107 pointed out, locomotion of
bacteria or other single-cell organisms in water occurs at low Reynolds number,107
where viscous drag dominates inertia, employing a geometric pumping mechanism
where only a non-reciprocal cycle of shapes can produce net motion.

In the early 1980s biochemist Tian Tsong,108 then at Johns Hopkins University,
began a series of experiments designed initially to study the phenomenon of elec-
troporation. In the course of his investigations with his co-workers,108 he made the
striking observation that AC electric fields applied to cells could cause the net trans-
port of ions from low to high electrochemical potential, i.e., in the non-spontaneous
direction. Detailed experiments showed86 that the observed behavior was indeed
mediated by the Na+,K+-ATPase and that the uphill transport was not simply a result
of activation of the pump’s ATP hydrolytic activity persisting even when there is no
possibility of ATP hydrolysis. The frequency response was explored in the early
1990s and could be interpreted86,109 in terms of relaxation kinetics.110 The

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Box 1. Trajectory Thermodynamics, Microscopic Reversibility, and Directional Cycling for Non-equilibrium Processes

MODEL FOR TRANSPORT ACROSS A MEMBRANE

(A) Model system with two energy levels that mediates transport of non-interacting particles between two reservoirs, left
(shown in green) and right (shown in blue), held at possibly different chemical potentials mL and mR , respectively. We assume
that the transporter can bind to only one particle at a time.(B) Triangle kinetic diagram99 for analyzing the net transport be-
tween the two reservoirs and the occupancy of the three possible states of the system, 0, 1, and 2. The transitions are color-
coded according to whether they are internal (red), involve particle exchange with the left reservoir (green), or particle ex-
change with the right reservoir (blue).

At equilibrium, when mL = mR , the ratio between any two-state concentrations is given by a Boltzmann expression, e.g.,


½2
½1 = eðG1 G2 Þ=RT , and there is no net flux between any two states, e.g., p½1eq  q½2eq = 0. When mL smR the situation is
eq


½2
more complicated. The concentration ratio ½1 is not in general given by a simple Boltzmann relation, and there can be
ss
net flux, J12 = p½1ss  q½2ss , between the states. We can analyze this non-equilibrium situation using ‘‘trajectory thermo-
dynamics,’’ a theory that has its roots in the work of Onsager and Machlup,100 and subsequent work of Terrell Hill,101
and that has recently formed the basis for a general theory of molecular motors presented by one of us.102 We start with
the thermodynamic constraints on the rate constant. The ratio between the internal transition constants p and q is
p ðG1 G2 Þ
= e RT (Equation 1)
q

and the ratios of the transition constants for exchange of particles between the system and the reservoirs are
a ðmL G1 Þ b ðG2 mR Þ k ðmR G1 Þ l ðG2 mL Þ
= e RT ; = e RT ; = e RT ; = e RT (Equation 2)
g d 4 h
To apply trajectory thermodynamic theory, we first focus on the ratio of the probabilities for clockwise versus counterclock-
wise cycling,
a+k p b+l
probð0 ! 1 / 2 
! 0Þ ða + kÞ ðb + lÞ p
3 = 3 (Equation 3)
d+h q
probð0 ! 2 / 1 ! 0Þ ðg + 4Þ ðd + hÞ q
g+4

There are four distinct cycles contained in the expression in Equation 3. Each cycle is constrained by microscopic revers-
a p b
ibility103 based on net effect of the cycle in the environment. For example, the cycle 0 / 1 / 2 / 0 involves transport of
d q g
a particle from the left reservoir to the right reservoir, and its microscopic reverse 0 / 2 / 1 / 0 involves transport of a par-
 
apb
ticle from the right reservoir to the left reservoir. The ratio of the product of rate constants is given by RT ln dqg = mL 

mR hDm. By use of Equation 2 the ratio in Equation 3 can be rewritten in terms of only Dm and the "off" rate constants, b,
l, g, and 4 as
2 g 3 2  3
Dm=RT
!0Þ 4 4 þ e þ1
aþk p bþl l
probð0!1/2 b
=  5 3 4l 5 (Equation 4)
dþh q
probð0!2/1!0Þ
gþ4 g
þ1 b
þ eDm=RT
4
|fflfflfflfflfflfflfflfflfflfflfflfflfflffl{zfflfflfflfflfflfflfflfflfflfflfflfflfflffl} |fflfflfflfflfflfflfflfflfflfflffl{zfflfflfflfflfflfflfflfflfflfflffl}
A1 A1
2

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where A1 and A2 are kinetic asymmetry factors introduced104 in the context of ATP hydrolysis-driven molecular motors.
Obviously if mR = mL then A1 A1
2 = 1, but for mR smL the directionality is controlled by the ratio of ‘‘off rate constants.’’ Taking
mR <mL , for 4g bl>1 then A1 A1 g b 1 g b 1 1
2 >1, for 4 l<1 then A1 A2 <1, and for 4 l = 1 then A1 A2 = 1. The ratio A1 A2 is independent of
G1 and G2. We can also use trajectory thermodynamics to calculate the steady-state ratios between the concentrations of
the three states, [0], [1], and [2]. For example, the steady-state between [1] and [2] occurs when the total probability of un-
dergoing a transition 1/2 is equal to the probability of undergoing the transition 2/1. For the cycle in Figure A there are
two paths (five trajectories S). One is the direct path with rate constants p and q and the other is the indirect path with prod-
ucts of the sums of rate constants ða + kÞðb + lÞ and ðg + 4Þðd + hÞ. Thus, the steady-state ratio is
  
½2 p + ½ðg + 4Þðd + hÞ  q + baeDm=RT + bk + la + lke + Dm=RT ðG1 G2 Þ=RT
= = e (Equation 5)
½1ss q + ½ðb + lÞða + kÞ  q + ba + bk + la + lk
|fflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl{zfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflfflffl}
W =RT 
e pd;S 1/2

where we used to write the ratio constants in the numerator in terms of those in the denominator reveal that the equilibrium
D E
term eðG1 G2 Þ=RT is modified by the term eW pd;S1/2 =RT is the exponential of the path dependent energy exchanged between
the reservoirs and the transporter averaged over all trajectories leading from state 1 to state 2. Note that the energy ex-
change obeys the symmetry W pd;S1/2 =  W pd;S 2/1 . This term, just like the the ratio A1 A1
2 , does not depend on the
free energies G1 or G2 . The response of the system to disequilibrium caused by being in contact with two reservoirs that
are not in equilibrium with one another depends only on the kinetics—the transition state energies—and not on the free-
energies of the states. From the way that Figure A is presented, it is very tempting to simplify the kinetic diagram by leaving
k h
off the transitions 0 # 1 and 0 # 2. This approach is thermodynamically unsound. The failure to show these transitions in the
4 l
kinetic diagram is equivalent to the implicit assumption that the coefficients k; 4;h;l = 0, which is obviously absurd—these
rate constants are tied to one another by the thermodynamic relations shown in Equation 2. The failure to include thermo-
dynamically necessary transitions is very common in the literature in many contexts, including for ATP-driven biomolecular
motors, and renders much of the published theory for "stochastic thermodynamics" of molecular machines suspect at
best.51

mechanism was termed111 ‘‘electroconformational coupling’’ in homage to Boyer’s

mechanism88—conformational coupling—for the FoF1-ATPase. These experiments
led to the theoretical realization that modulation of the energies of kinetic barriers
and binding sites could provide a mechanism for the controlled input of energy
into a wide range of molecular systems in order to drive them into a variety of
non-equilibrium steady states by what has come to be known as an energy ratchet
mechanism,63,64 an example of which is given in Figure 2D.

Figures 2D and 2E illustrate how the idea of geometric pumping can be used in
quantification of AC electrical field pumping mediated by Na+,K+-ATPase. For
simplicity, we conceptualize the pump as moving only a single ion per cycle. In
this case, a pump can be thought of energetically as a transmembrane protein
with parts that present barriers to an ion on both sides of a membrane with a binding
site between the two barriers. This sequence of a barrier, a binding site, and a barrier
separating two reservoirs forms what we term a pumping cassette. By controlling the
heights of the barriers, the access for ions from the two sides can be regulated, and
by modulating the energy of the binding site, the probability that an ion is bound is
controlled. The energy landscape for an ion to cross the membrane through a pump
protein can be described in terms of the relative barrier heights u, and the well
depth ε. At the bottom of Figure 2D, there is a kinetic scheme describing the
transport process with rate constants that can be expressed in terms of u and ε.
Remarkably, as shown in Figure 2E, when an AC electric field is applied to

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erythrocytes, what occurs86 is pumping of Rb+, an analog of K+ that has a radioactive

isotope, and Na+ ions from low to high electrochemical potentials without the
hydrolysis of ATP. The energy comes from the applied field, and for mL zmR
the frequency response of the ion flux is characterized by a single expression,
u ~2
~ + Anonad u
jIL/R jz , where u ~ is the frequency of the applied field divided by the
1+u ~2
relaxation frequency (1 kHz for Rb+ and 1 MHz for Na+) of the transport processes.
Note that if either ε or if u is time-independent, the pumped current is zero.112

We can also express the steady-state chemical potential difference after a long time
 
(many cycles of the field) as eðmR mL Þ=RT ss = eW pd;SðL/RÞ where W pd;SðL/RÞ is the energy
exchanged between the pump and the environment in a specific trajectory SðL/RÞ

for an ion to move through the pump protein, and eW pd;SðL/RÞ is the exponential of
this work averaged over all possible trajectories for the passage of an ion from the
left reservoir to the right reservoir. This exponential average, which depends on
the relationship between the modulation of the well energy, εðtÞ, and the relative
barrier heights, uðtÞ, is not a state function113 and can be either greater
than or less than 1. For more discussion on trajectory thermodynamics,
see Box 1.99–104

Inspired by the pumping-cassette structure evident in Figures 2C and 2D with a well,

the energy of which can be modulated, surrounded by two barriers whose relative
energies can also be modulated, we began an exploration of the small-molecule im-
plementation of this concept in 2013 using114 an artificial supramolecular assembly
(Figure 3A) that can undergo redox-driven unidirectional relative translational mo-
tion, employing a flashing energy-ratchet mechanism.63,64 The design principle re-
lies solely on the switchable noncovalent bonding interactions that simultaneously
regulate (Figure 3A) two kinetic barriers and one thermodynamic well. The constitu-
tionally asymmetric pseudodumbbell (PDB+) consists of a centrally located electron-
rich 1,5-dioxynaphthalene (DNP) recognition site for the electron-deficient cyclobis(-
paraquat-p-phenylene) (CBPQT4+) ring, a neutral 2-isopropylphenyl (2IPP) group at
one end and a positively charged 3,5-dimethylpyridinium (3/5PY+) unit at the other
end. The unidirectional movement of the ring along and off PDB+, involving the
complexation and de-complexation of a pseudo[2]rotaxane, is realized by modu-
lating the redox state of the ring between CBPQT4+ and CBPQT2(,+). In its fully
oxidized state, the ring undergoes thermal activation over the 2IPP unit to the
DNP recognition site. Upon reduction, the stability of the complex is significantly
decreased and those rings that dissociate do so overwhelmingly over the 3/5PY+
unit. The movement onto the pseudodumbbell over the 2IPP unit and off PDB+
over the 3/5PY+ unit is the non-reciprocal process necessary for the geometric
pumping of unidirectional motion. Similar directional processes have been demon-
strated114 using chemical and electrochemical fuels as well as by light.

In 2015, Credi and co-workers61 described an example of a self-assembling and dis-

assembling supramolecular system capable of relative unidirectional transport of a
ring across an asymmetric pseudodumbbell using light as the only energy source.
The key to the directionality lies (Figure 3B) in an azobenzene photoisomerization
that is capable of controlling both the noncovalent bonding interactions (thermody-
namic wells) and kinetic barriers for threading and de-threading over both ends of
the asymmetric pseudodumbbell (E)-ADB+, which consists of a central ammonium
ion recognition site115 for a 2,3-dinaphtho[24]crown-8 ether DN[24]C8, a photo-
switchable (E)-azobenzene unit at one end, and a methylcyclopentyl steric barrier

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Figure 3. Unidirectional Molecular Transport

(A) Graphical representations (top) of the structural formulas for the CBPQT 4+ ring and the PDB +
pseudodumbbell, which consists of a centrally located electron-rich DNP recognition site for the
electron-deficient CBPQT 4+ ring, a neutral 2IPP unit at one end and a positively charged 3/5PY+
unit at the other end. The PF 6 – counterions are omitted for the sake of clarity. The unidirectional
threading and de-threading along the pseudodumbbell PDB + (middle) are realized by altering the
redox state of the CBPQT4+ ring. At the outset, the CBPQT4+ ring binds to the DNP unit most likely
by passage over the 2IPP unit rather than over the positively charged 3/5PY + unit on account of
electrostatic interactions. Subsequent reduction of CBPQT 4+ to CBPQT (,+)(2+)/CBPQT2(,+)
diminishes the Coulombic repulsions between the reduced ring and the 3/5PY + unit. The reduced
ring most probably slips off the pseudodumbbell by passing over the 3/5PY+ unit, which presents a
smaller barrier to the ring than does the 2IPP unit. Upon oxidation, the ring reverts to its
tetracationic state, thus resetting the system. Energy profiles (bottom) represent the free energies

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Figure 3. Continued
of the system as the ring is transported along and off the pseudodumbbell unidirectionally via a
pseudo[2]rotaxane. Adapted with permission from Li et al. 114 Copyright 2013 American Chemical
Society.
(B) Graphical representations (top) of the structural formulas for the ring DN[24]C8 and the
pseudodumbbell ADB + in two isomeric forms (E and Z), which consists of a central ammonium ion
recognition site for the ring, a photoswitchable azobenzene unit at one end and a
methylcyclopentyl steric barrier at the other end. The light-driven unidirectional molecular
transport is enabled by the photoswitchability of the azobenzene unit powered solely by light.
Energy profiles representing the free energies of the system as the ring is transported along and off
the pseudodumbbell unidirectionally, via the intermediacy of pseudo[2]rotaxane, are illustrated at
the bottom. Adapted with permission from Ragazzon et al. 61 Copyright 2015 Springer Nature.

(pseudo-stopper) at the other end. The crown ether threads onto the pseudodumb-
bell and encircles the ammonium-ion recognition site as a result of hydrogen-
bonding interactions, augmented by [p–p] stacking interactions between the naph-
tho and azobenzene units, which occurs preferably from the (E)-azobenzene side of
the pseudodumbbell on account of a relatively small kinetic barrier compared with
that of the neutral pseudo-stopper. Subsequently, light irradiation triggers efficient
photoisomerization of the azobenzene unit from its (E)- to its bulkier (Z)-form, thus
diminishing the hydrogen-bonding interaction between the ring and the pseudo-
dumbbell (Z)-ADB+ accompanied by an enhanced steric barrier arising from the re-
sulting (Z)-azobenzene unit. These changes in the potential energy landscape cause
the ring to traverse the methylcyclopentyl steric barrier and de-thread unidirection-
ally from the other end of the pseudodumbbell. The reset of the system is achieved
by the conversion of the (Z)-azobenzene unit back to its original (E)-isomer under
either irradiation or thermal relaxation. The prerequisite for the system to operate
continuously and autonomously is the overlapping absorption spectra of (E)- and
(Z)-azobenzene since both (E) to (Z) and (Z) to (E) photoisomerization can happen un-
der the same photochemical conditions. In order to prove the working mechanism of
this supramolecular ensemble, the authors carried out many detailed experiments
on the system using 1H nuclear magnetic resonance (NMR) spectroscopy, UV-vis ab-
sorption spectrophotometry, luminescence spectroscopy, and theoretical calcula-
tions. The data they obtained provide convincing evidence to support the conclu-
sion that the self-assembling ensemble operates directionally by consuming light
energy. This supramolecular system can be looked upon as a prototypical light-
driven AMP.

ARTIFICIAL MOLECULAR PUMPS

Regardless of the mechanistic differences between pumps, either those powered by
human hands in the ancient macroscopic world or ATP hydrolysis in the microscopic
biological world, the ability for them to orchestrate directional mechanical motion
involving components is essential to produce useful work and perform specific tasks.
In stark contrast, synthetic chemists have faced a long-standing challenge of
designing and creating molecules capable of biasing conformational and co-confor-
mational changes—in other words, molecules possessing movable components
with controlled motions. As a result, substantial progress in the field of AMMs has
been accompanied largely by the development of effective synthetic approaches
to MIMs, such as rotaxanes and catenanes, during the past 35 years.116 The applica-
tion of template-directed synthesis117–120 and dynamic covalent chemistry121 has
contributed to the construction of (supra)molecular entities that can perform simple
tasks and, to some degree, resemble their biological counterparts. From here on, we
will discuss several examples of AMPs reported to date from our own research

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laboratory42,57–59 as well as from David Leigh’s laboratory.60 The discussion will be

organized according to the underlying design and operation governing these mo-
lecular machines and the types of stimuli controlling the relative directional move-
ments of their components.

AMPs based on MIMs operate in an environment dominated by random thermal fluc-

tuations, i.e., Brownian motion.72 These AMPs use external redox energy to rectify
this random motion through ratcheting mechanisms,51 which are also widely consid-
ered to be the principles by which the motor proteins operate in the biological
world.122

In 2010, we reported123 an example of radically enhanced molecular recognition

that represents an enormously valuable tool for the design and synthesis of
AMMs. Under reducing conditions, the bipyridinium radical cation (BIPY+,) is able
to form an extremely strong inclusion complex (Ka = 7.9 3 104 M–1 determined124
in MeCN by UV-vis spectrophotometry) with the bisradical dicationic cyclobis(para-
quat-p-phenylene) (CBPQT2(+,)), forming a pseudo[2]rotaxane on account of radical-
radical interactions.125 Later in the same year, this newly discovered noncovalent
bonding interaction was put126 to work in the template-directed synthesis of a [2]ro-
taxane,117–120 containing only electron-deficient CBPQT4+ and BIPY2+, representing
a type of MIMs composed of components with little or no binding affinities in their
ground states. Since then, the radical templation protocol has become widely
used127 in template-directed synthesis to obtain a range of catenanes with densely
packed positive charges. For example, a highly energetic octacationic homo[2]cat-
enane composed of two CBPQT4+ rings, which was reported128 by us in 2013, exists
as a persistent monoradical septacation that is stable under ambient conditions.
Employing the click reaction,129,130 we reported131 recently the template-
directed117–120 synthesis of a dodecacationic [3]catenane consisting of three me-
chanically interlocked 4+ charged rings—two of them are CBPQT4+—and a tetraco-
sacationic radial [5]catenane, consisting of four CBPQT4+ rings mechanically inter-
locked around a larger 8+ charged ring, bearing a total of 24 positive charges in
its co-constitution. The dodecacationic [3]catenane was found to possess the high-
est charge density ever observed in a MIM. The use of redox-switchable noncovalent
bonding interactions based on radical dimerization has been key to the design and
synthesis of a variety of molecular switches and machines based on rotaxanes132,133
and catenanes134—especially relating to the development of redox-driven AMPs
that will now be discussed in detail.

The design and synthesis of a redox-driven AMP with the ability to concentrate posi-
tively charged rings onto a short collecting chain on a dumbbell, creating highly non-
equilibrium rotaxanes, was a signal achievement.42 The unidirectional pumping of
several rings from the bulk solution onto the dumbbell relies on a flashing energy-
ratchet mechanism,63,64 resulting from the modulation of kinetic barriers and ther-
modynamic wells.

In detail, the unique feature of the first generation (Mark I) of AMPs lies in the design
of the dumbbell 1-MP3+, illustrated in Figure 4A. This dumbbell-shaped molecular
pump consists of (1) a redox-active bipyridinium (BIPY2+) unit positioned between
(2) a 3/5PY+ Coulombic barrier and (3) an isopropylphenylene (IPP) steric barrier,
and (4) a short oligomethylene collecting chain, comprising of 11 carbon atoms,
terminated by (5) a bulky 2,6-diisopropylphenyl stopper (S). Operating in a fashion
similar to membrane transport proteins in nature and using radical-pairing interac-
tions, the pump possesses exquisite structural features that allow for control over

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Figure 4. The First Artificial Molecular Pump (Mark I)

(A) Graphical representations of the structural formulas of CBPQT 4+ and 1-MP 3+, which consists of
a redox-active BIPY 2+ unit positioned between a 3/5PY + Coulombic barrier and an IPP steric barrier,
and a short oligomethylene (containing 11 methylene groups) collecting chain terminated with a
bulky stopper (S). The PF 6 – counterions are omitted for the sake of clarity.
(B) The operation of the Mark I 1-MP 3+ in the presence of the CBPQT 4+ ring is triggered by
repetitive reduction and oxidation, as well as by thermal activation, leading to the production of a
[2]rotaxane and then a [3]rotaxane. At the outset, the rings and the dumbbell repel (BI) one another
on account of strong Coulombic repulsions. Upon reduction, all BIPY 2+ units are reduced to their
radical cationic (BIPY+,) states, leading to the threading (BII) of a CBPQT 2(+,) ring, followed by the
formation of a trisradical tricationic complex. On oxidation, the renewed strong Coulombic
repulsions between the charged 3/5PY + , BIPY2+ and CBPQT 4+ units cause the ring to occupy a
metastable state prior to traversing (BIII) the IPP steric barrier and falling, as a result of thermal
energy, into a kinetic trap provided by the oligomethylene collecting chain, resulting in the

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Figure 4. Continued
formation (BIV) of a [2]rotaxane. The second reduction allows the pump to recruit (BV) another ring
from the bulk solution following a similar mechanism. The second oxidation restores full charges to
the BIPY2+ units and the CBPQT 4+ rings and obliges the second ring to traverse (BVI) the IPP steric
barrier. Thermal relaxation results in the formation (BVII) of a [3]rotaxane. Profiles representing the
free energies of the system as a ring traverses the dumbbell are illustrated to the right of each
intermediate in the reaction sequence. The curved arrows on the energy profiles represent reaction
pathways that are either kinetically favored (green) or disfavored (red). The steric barrier imposed
by IPP remains the same throughout the redox cycle.

a series of co-conformational rearrangements along a kinetically favorable pathway

that allows two rings to move sequentially away from equilibrium toward a higher
local concentration on the collecting chain of the dumbbell.

At the outset, the molecular pump 1-MP3+ and the CBPQT4+ ring repel (Figure 4BI)
one another on account of strong repulsive Coulombic interactions. Upon addition
of an excess of Zn dust, the bipyridinium units present in both the ring and on the
pump are reduced to BIPY+, units, leading to radical-pairing between CBPQT2(+,)
and BIPY+,, forming (Figure 4BII) a stable trisradical tricationic complex. Subsequent
oxidation restores three positive charges to the pumping system, bringing the total
charge to +7 and obliging the ring to traverse (Figure 4BIII) the IPP steric barrier with
the aid of thermal energy augmented by the accumulative effect of the electrostatic
barriers imposed by both the 3/5PY+ and BIPY2+ units on the pumping cassette. A
series of co-conformational changes accompany the translocation of the ring over
the IPP unit onto the oligomethylene collecting chain, affording (Figure 4BIV) a kinet-
ically stable out-of-equilibrium [2]rotaxane. The key to the successful recruiting of a
second ring onto the pump, following a further reduction, resides in the steric hin-
drance imposed by the IPP unit and the lack of Coulombic repulsions between
related component parts. It is kinetically more favorable to recruit (Figure 4BV) a sec-
ond reduced ring from the bulk solution than from the collecting chain. Subsequent
oxidation induces the passage (Figure 4BVI) of the oxidized ring encircling the
BIPY2+ unit over the IPP unit to join the first ring. The resulting [3]rotaxane (Fig-
ure 4BVII) with 11 positive charges is highly energetic but kinetically very stable on
account of the large barrier imposed by the IPP unit.

In terms of the pumping energetics, the redox operation of the AMP relies on a
flashing energy-ratchet mechanism,63,64 where the redox-driven modulation of the
energy profiles (Figure 4B) biases the Brownian movement of each ring to favor an
accumulation of two rings on the collecting chain. Symmetry breaking113,135 is the
key to the operation of the energy-ratchet mechanism.63,64

Such subtle and delicate molecular design has proven to be successful only after
much trial and error, aided and abetted by extensive computational and experi-
mental screening.136 It should be noted that the lengths of the linkers between (1)
the 3/5PY+ and BIPY2+ units, (2) the BIPY2+ and IPP units, and (3) the IPP and S units,
all play a critical role in determining kinetic barriers and thermodynamic wells that
influence the pumping reliability in a highly sensitive manner. Kinetic measurements
by 1H NMR spectroscopy in CD3CN and subsequent analyses reveal all but identical
activation energies (DGs) required for the first (23.0 kcal mol1) and second
(23.1 kcal mol1) rings to pass over the IPP steric barrier during successive indepen-
dent redox cycles. This observation tells us that the presence of the first ring on the
collecting chain does not inhibit the passage of the second ring to join it. The static
transition state associated with the IPP steric barrier is an important property going
forward.

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Figure 5. Mark II Molecular Pump

Graphical representations (top) of the structural formulas of CBPQT 4+ and 2-MP 3+, which consists
of a redox-active BIPY 2+ unit positioned between a 2/6PY + Coulombic barrier and an IPP steric
barrier, and a short oligomethylene (containing 12 methylene groups) collecting chain terminated
with a bulky stopper (S). The PF 6 – counterions are omitted for the sake of clarity. The operation
(bottom) of the Mark II 2-MP 3+ in the presence of the CBPQT4+ rings is triggered by repetitive
reduction and oxidation, as well as by thermal activation, leading to the production of a [2]rotaxane
and then a [3]rotaxane. The operation of this Mark II pump is rapid as a result of the enhanced
electrostatic repulsion between BIPY2+ and CBPQT4+ as a consequence of the constitutional
modifications 1 (the substitution of 3/5PY + by 2/6PY +) and 2 (the reduction of the length of the
spacer between the BIPY 2+ and the IPP unit from two carbons to one). Adapted with permission
from Pezzato et al. 59 Copyright 2017 Elsevier.

With all these design principles well established, we reported59 (Figure 5) a second-
generation (Mark II) of AMP in which the 3/5PY+ was substituted (Figure 5, modifica-
tion 1) by a 2,6-dimethylpyridinium (2/6PY+) Coulombic barrier and the reduction
(Figure 5, modification 2) of the length of the spacer between the BIPY2+ and the
IPP unit from two carbons to one. These minor modifications, which enhance the
electrostatic repulsion between the BIPY2+ unit on the dumbbell and the CBPQT4+
ring, lower considerably the activation energy (21.8 kcal mol1) for the passage of

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the ring over the IPP steric barrier, thus making it possible for the Mark II pump to
operate more rapidly than the Mark I pump.

The use of chemical reagents is intrinsically problematic because of the need for re-
petitive additions of both reductants and oxidants, leading to the accumulation of
waste products. This problem was addressed in a report published58 in 2018 from
our laboratory, describing a different approach to operating a duet molecular
pump away from equilibrium. Two Mark II AMPs were attached covalently in a tail-
to-tail manner to both ends of a short oligomethylene collecting chain incorporating
two dimethyl ammonium ion centers. The CBPQT4+ rings were added in pairs to the
collecting chain in a controlled manner using a home-built electrochemical cell. This
electrochemical approach allows the duet molecular pump to entice a pair of rings
from the bulk solution onto the collecting chain, conducting each redox cycle by
means of a controlled supply of electricity, leading to the formation of a [3]rotaxane
and then a [5]rotaxane after two redox cycles.

In late 2019, we extended57 the scope of the redox-driven AMPs with respect to both
their designs and functions. In the event, a molecular dual pump (MDP6+) was pre-
pared (Figure 6) by linking two Mark II AMPs together covalently in a head-to-tail
manner, forming a collecting oligomethylene chain in the middle between the two
pumps. Two sequential molecular pumping events, powered either by redox re-
agents (Zn and NOPF6) or by alternating redox potential (0.7 and +1.4 V), led to
the assembly and disassembly of an intermediate [2]rotaxane, wherein the CBPQT4+
ring undergoes linear unidirectional motion along the molecular dual-pump dumb-
bell. The redox operation of this dual pump, which can be driven either chemically or
electrochemically using an energy-ratchet mechanism,63,64 allows for the controlled
capture and release of a ring from and, subsequently, back into the bulk solution.
One design feature of this dual pump is that it provides potential access to the inte-
gration of a functionalized pump with itself or other types of AMMs conveniently by
using click chemistry.129,130 The dual pump may be capable of directional transport
of substrates over long distances.

In summary, the design and synthesis of a range of redox-driven AMPs have led us to
the preparation of enthalpically and entropically demanding rotaxanes as a result of
ratchet-driven unidirectional transport. By recruiting multiple rings away from equi-
librium, these wholly synthetic AMPs create a concentration gradient in an artificial
system, serving as a minimalistic design to perform tasks just as transport proteins
do in living organisms.

In 2017, Leigh and colleagues60 described two chemically driven artificial rotary mo-
tors and one linear molecular pump using a similar molecular design, all three of
which are capable of conducting unidirectional molecular motion. The use of a
chemical reagent (trichloroacetic acid) as the sole fuel, added in pulses, powers
the operation of the motors and pump following an energy-ratchet mechanism63,64
by controlling simultaneously the binding affinities (thermodynamic wells) and the
relative heights of acid-base labile steric barriers (kinetic barriers). Here, we focus
our attention on the design and operation of the chemically fueled AMP illustrated
in Figure 7. The chemically driven symmetrical dumbbell CDB comprises (Figure 7)
two working pumps, attached together in a tail-to-tail fashion by a long oligomethy-
lene chain, which, however, does not serve as a collecting chain like the previously
described redox-driven AMPs. Each individual pump consists of two recognition
sites, including an acid-base switchable center involving a secondary ammonium
ion recognition site and a secondary amine (recognition switched off) and an inert

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Figure 6. A Molecular Dual Pump

The pumping mechanism for controlled molecular transport (top) by a dual pump. The ring and the
dumbbell repel (I) one another initially on account of strong Coulombic repulsions. Reduction by Zn
dust, or a constant negative potential (0.7 V), favors the formation (II) of two thermodynamically
stable trisradical tricationic complexes. Oxidation by NOPF 6 , or a constant positive potential (+1.4
V), restores Coulombic repulsion between the ring and the BIPY2+ unit on the dumbbell causing the
first ring to pass over the steric barrier (IPP) by thermal relaxation and threading onto the collecting
chain, thus forming (III) an isolable [2]rotaxane, while the second ring departs the dumbbell and
returns into the bulk solution. Another reduction performed either chemically or electrochemically
on the isolated [2]rotaxane, with no free CBPQT 4+ in the solution, triggers the complexation (IV) of
the threaded ring with the second pump by dint of a kinetically favored process. The subsequent
oxidation obliges the ring to depart from the dumbbell and resets the system. Profiles representing
the free energies of the system as the ring traverses along and off the dumbbell via a stable [2]
rotaxane intermediate are illustrated at the bottom. The curved arrows are representations of
reaction pathways that are kinetically favored (green) or disfavored (red). The dashed black lines
indicate the redox-independent steric barrier imposed by IPP. Adapted with permission from Qiu
et al. 57 Copyright 2019 American Chemical Society.

triazolium ring, and two steric barriers, based on bulky hydrazones and disulfides
with opposite labilities in acids and bases. The addition of trichloroacetic acid to
the system containing the crown ether rings DB30C10 and the dumbbell CDB re-
moves the two acid-labile hydrazone blocking groups and protonates the dibenzyl-
amine centers to form the corresponding ammonium ion recognition sites for the
rings, leading to pseudo[3]rotaxane formation. It should be noted that the rings

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Figure 7. A Chemical-Pulse-Driven Molecular Pump

The chemically driven operation of the dumbbell CDB, consisting of two working pumps attached
together by an oligomethylene chain, with pulses of chemical fuel to afford [3]rotaxane 3R and [5]
rotaxane 5R by sequestering the dibenzo-30-crown-10 DB30C10 in a solution of CD2 Cl 2 :CD 3 CN
(9:1). Each individual pump consists of two recognition sites, including a switchable center
involving a secondary ammonium ion recognition site and a secondary amine (recognition switched
off) and a triazolium ring, and two steric barriers, based on hydrazones and disulfides with opposite
labilities in acids and bases. The addition of one pulse of fuel (trichloroacetic acid) removes the two
acid-labile hydrazone groups and protonates the two dibenzylamine centers to form the
corresponding ammonium ion recognition sites for the two rings, leading to pseudo[3]rotaxane
formation. The presence of a base (triethylamine) catalyzes the transformation of trichloroacetic
acid into chloroform and carbon dioxide, resulting in the whole system becoming basic, thus
triggering (1) reattachment of the hydrazone groups, (2) deprotonation of the ammonium ion
recognition sites to afford the dibenzylamines, and (3) removal of the bulky disulfide groups. As a
result, the rings thread onto the inner triazolium recognition sites, forming a stable [3]rotaxane 3R.
An additional three pulses of acid results in the formation of [5]rotaxane 5R as the major product of
this energy-ratchet mechanism. 63,64 Adapted with permission from Erbas-Cakmak et al.60
Copyright 2017 American Association for the Advancement of Science.

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do not bind to the inner triazolium rings since the pathways to allow this interaction
to happen are blocked by the disulfides units under acidic conditions. The ingenuity
of the design comes in what happens next. In the presence of a base (triethylamine),
which serves as a poor catalyst to break down trichloroacetic acid into chloroform
and carbon dioxide, the whole system gradually becomes basic as the chemical
fuel (trichloroacetic acid) disappears, thereby triggering (1) reattachment of the
bulky hydrazone blocking groups, (2) deprotonation of the ammonium ion recogni-
tion sites to afford the dibenzylamines, and (3) removal of the bulky disulfides group.
As a result, the mechanically interlocked rings eventually thread over onto the inner
triazolium recognition sites, forming a stable [3]rotaxane 3R following a single pulse
of fuel. An additional three pulses of the acid into the system results in the formation
of a [5]rotaxane 5R as the major product by means of an energy-ratchet mecha-
nism.63,64 Overall, these operationally simple and efficient wholly synthetic molecu-
lar motors and pump, powered by pulses of a chemical fuel, represent a significant
advance toward controlling the directional movement of substrates at the nano-
scopic level.

EXPLOITING CATALYSIS IN NON-EQUILIBRIUM CHEMISTRY

Most biomolecular motors and pumps use catalysis of an energy-releasing (exer-
gonic) chemical reaction to power directed motion and/or thermodynamically uphill
pumping.104 Often, the reaction is ATP hydrolysis, which, under normal physiolog-
ical conditions, releases about 20 RT energy per mole hydrolyzed. Recently, Wilson
et al.137 have reported the first example of a catalysis-driven synthetic rotor, shown
schematically in Figure 8. A small benzylic amide ring—illustrated in the cartoons in
dark blue—undergoes (Figure 8B) directional rotation around a large loop. In this [2]
catenane two fumaramide recognition sites—shown in aqua in the cartoons—
interact with the blue ring. Directionality is conferred on the circumrotation of this
ring between the two recognition sites on the loop by the timely addition and
removal of bulky protecting groups. These groups are added by catalytic esterifica-
tion with fluoromethoxycarbonyl chloride (Fmoc-Cl), shown in the cartoons in pink.
Significantly, removal of the protecting group in the form of dibenzofulvene
(DBFV) occurs spontaneously by a non-catalytic cleavage as shown in Figure 8C.
There are three relevant co-conformations of the [2]catenane differing in the location
of the blue ring relative to the pink protecting groups. In one co-conformation, Fum-
1, both hydroxyl groups (red) are esterified with Fmoc. In another co-conformation,
Fum-2, the blue ring is located near the Fmoc protecting group, while in the third co-
conformation, Fum-20 , the blue ring is distant from the protecting group. The con-
centration of reagents was adjusted such that only one hydroxyl group at most
was exposed at any one time. The net outcome of derivatizing and removing the pro-
tecting group is the conversion of Fmoc to DBFV. This conversion is exergonic and,
under the experimental conditions, releases about 25 RT energy, roughly compara-
ble to the energy released by ATP hydrolysis.

There are two important factors to consider regarding the directionality of this mo-
lecular rotor. Firstly, the equilibrium, Fum  2#Fum  20 is shifted to the right. This
shift makes a prima facia case for the blue ring rotating clockwise, in the order Fum-1
/ Fum-2 / Fum-20 / Fum-1. There is, however, a kinetic effect. The catalytic
esterification to with Fmoc-Cl is faster in Fum-2 than in Fum-20 . This effect argues
in favor of counterclockwise circumrotation of the blue ring in the order Fum-1 /
Fum-20 / Fum-2 / Fum-1. To determine which way the ring moves with respect
to the loop, we must carry out a quantitative analysis using the constraints of micro-
scopic reversibility.103 When this analysis is carried out, the ratio, r, for the relative

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Figure 8. Catalysis-Driven Synthetic Molecular Rotor

(A) Graphical representations of the structural formulas of the [2]catenane Fum-1. Under the
experimental conditions, the [2]catenane has three distinct chemical states. The state with both
hydroxyl groups unprotected has a very low concentration given the amount of Fmoc-Cl that was
used in the experiment. Adapted with permission from Wilson et al. 137 Copyright 2016 Springer
Nature.
(B) The kinetic mechanism for interconversion between the three states Fum-1, Fum-2, and Fum-2 0 .
There are two effects to consider, the thermodynamic shift of the equilibrium between Fum-2 and
Fum-2 0 in favor of the latter and the kinetic difference in the rate of catalytic protection by Fmoc. In
order to analyze which of these effects determine the directionality, it is necessary to solve kinetic
equations described in the text, using the constraints of microscopic reversibility
!
kFmoc DBFV
on;i koff;i
Dm = RT ln kFmoc DBFV for i = 2;20 , where Dm is the free-energy released in the conversion of Fmoc-Cl
off;i kon;i

to DBFV. The results are discussed in the text.

(C) The mechanism for conversion of Fmoc-Cl to DBFV at the two hydroxyl groups on the large loop.

likelihood of the blue ring to undergo clockwise circumrotation pð1/2/20 /1Þ

versus counterclockwise circumrotation pð1/20 /2/1Þ is found to be

! !
kFoff;20 Dm=RT kFoff;2 F
+e +1 kFoff;2 koff;20 kF 0
kD
off;20
kD
off;2 kD D + koff;2
D
off;2 koff;20 off;20
r= ! !z F
kF kFoff;2 koff;20 kF
off;2
0
+1
kFoff;2
+ eDm=RT kD D + koff;2
D
kD 0 kD off;2 koff;20 off;2
off;2 off;2

where the approximation is based on observation that Dmz25RT, the energy

released by the conversion of Fmoc to DBFV, is very large and hence the exponen-
kF kF 0
tials can be neglected in comparison with the ratios koff;2
D and koff;2
D . Note that the ratio
off;2 off;20

Chem 6, 1952–1977, August 6, 2020 1971

 ll
Review

k2;20
k20 ;2 plays no role whatsoever in determining the directionality of the blue ring. Since

kD D Fmoc Fmoc
off;2 zkoff;20 and koff;2 >koff;20 , the ratio r is less than 1, i.e., the ring undergoes coun-
terclockwise circumrotation, as observed experimentally. The analysis carried out
here for the specific small-molecule motor reported by Wilson et al.137 is general
and highlights an important point. The design principle for catalysis-driven pumps
and motors is very different from that for pumps and motors driven by external mod-
ulation. Externally driven AMPs (Figures 3, 4, 5, 6, and 7) require that the energy of
the recognition sites must change as a function of the modulation. In stark contrast to
these cases of externally driven molecular machines, modulation of the energy of the
recognition sites is unimportant for those driven by catalysis, where the key factor is
gating the catalysis through allosteric interactions by which an information ratchet135
is driven.

OUTLOOK
We have highlighted a selection of macroscopic, biological, and AMPs through the
ages with the emphasis being placed on their design and underlying operating prin-
ciples. We have also devised a theoretical framework to facilitate the design of arti-
ficial molecular motors, including pumps, driven by catalysis of a chemical reaction
employing an information ratchet.64,135

The past few decades have witnessed substantial advances in the field of AMMs in
the wake of advances in unnatural product synthesis.50 Our ability to control the rela-
tive motion of molecular components in a nanoscopic world has led us to the design
and synthesis of a plethora of AMMs that operate away from equilibrium. It remains,
however, a significant challenge for researchers in this field to harness the controlled
motions of AMMs to organize collectively and perform work on their macroscopic
surroundings. One might ask, what have we achieved with the design and synthesis
of redox-driven AMPs? The answer is that the combination of each individual func-
tional component (e.g., PY+, BIPY2+, IPP, and CBPQT4+) working in concert has
led to emergent machine-like behavior during the transport of rings uphill onto col-
lecting chains iteratively and progressively against local chemical gradients. The
integration of multiple AMMs, where the upstream controlled molecular motion
and function is transcribed into signals for the downstream molecular machines,
leads to machines with ever-increasing sophistication and potentially emergent
properties. The recent advent of systems chemistry16–18,138–142 is set to play a vital
role in realizing this goal through the design of complex chemical systems net-
worked in a highly programmable manner while introducing feedback loops similar
to those in systems biology.143

The immediate exploitation of redox-driven AMPs could be to transport substrates

(rings) between different reservoirs, creating concentration gradients. Possible im-
plementations include the incorporation of such pumps into bilayer membranes
and their immobilization onto surfaces144–146 or inside frameworks.147 Research on
this topic is currently underway in our laboratory. The recent development of nano-
confinement technologies,148 such as nanopores149,150 and nanopipettes,151 may
also provide a useful platform for controlling and monitoring the relative motions
of the components of AMMs. Recently, we have also witnessed a renaissance of
research into the use of MIMs in sensing152 and catalysis153 by taking advantage
of the crowded environments created during the formation of mechanical bonds.
Recently, Goldup and co-workers154 reported the first example of enantioselective
catalysis with a mechanically planar chiral rotaxane. We find it intriguing that most

1972 Chem 6, 1952–1977, August 6, 2020

 ll
Review

of the previous syntheses of AMPs and molecular motors have focused on designing
sophisticated dumbbells (energy surfaces) while the rings (Brownian particles)
remain relatively simple and symmetrical. The introduction of symmetry
breaking113,135 and chiral elements155 into the design of rings may hold potential
for the development of new molecular machines.

Another potential use of AMPs is found in the precise synthesis of polyrotax-

anes.156 Chemically or electrochemically controlled redox reactions provide
ready access to polyrotaxanes with a prescribed number of rings threaded
onto polymer dumbbells, irrespective of the strength of the noncovalent
bonding interactions between the polymer and the rings. What could come
next? What about a highly efficient polyrotaxane synthesizer,157 which can be
produced by attaching two AMPs to both ends of a polymer collecting chain?
It should be possible to generate highly engineered polyrotaxanes, with palin-
dromic arrays of co-constitutionally heterotopic rings positioned on constitution-
ally symmetrical polymer dumbbells and then ultimately to transcribe their pro-
grammed information back into the domain of sequence-controlled polymer
synthesis.158

Where else can we go in search of designs for artificial pumps and motors that rely on
mechanical bonding? One answer lies in the rapidly developing field159–161 of mo-
lecular nanotopology,162 presently fueled by a rapid increase in the synthesis of arti-
ficial trefoil163–168 and pentafoil knots.169–171 Nature—both in DNA and proteins—
provides examples172 of knotted molecules, presumably because knotting is a
way of decreasing a molecule’s degree of freedom, as well as introducing tight-
ness173–175 and nanoconfinement148 on them even in an aqueous solution.176 The
presence of knots in molecules limits the number of conformations chains can adopt
and may also allow the formation of active sites, modulation of which may be
capable of driving chemical reactions. Artificial molecular knots are worthy of atten-
tion by molecular machinists177–179 in designing and synthesizing molecular
machines.180,181

The combination of energy and information ratchets63,64 in the operation of artificial

molecular motors may provide a beneficial and promising way to design the next
generation182 of AMMs and achieve some of the sophistication found in nature’s bio-
logical pumps. In turn, the design and synthesis of these machines from the bottom-
up can also serve as a multi-disciplinary entry point to understanding how transport
proteins work, both in terms of the importance of allosteric mechanisms of gating
catalysis183–185 and for understanding the importance of the aqueous solvent in
which they carry out their functions.186 It is our calling as scientists to create the un-
natural molecular counterparts to biomolecular machines to help unravel the origin
of life.53 We are reminded of the phrase on Feynman’s last blackboard, ‘‘What I
cannot create I do not understand.’’

ACKNOWLEDGMENTS
The authors would like to thank Northwestern University for its continuing financial
support.

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