USING PROTEIN CHEMISTRY AS A WAY INTO TEACHING CHEMICAL

BONDING TO A-LEVEL CHEMISTRY STUDENTS

Introduction
I doubt that there can be many experienced chemistry teachers who have not felt a
certain sense of ennui, knowing that they have to start the A-level course in September with
the inevitable ‘structure and bonding’ topic. There isn’t a lot of practical work to do, the topic
is abstract and difficult and, let’s face it, pretty dry too. However, I have discovered a
different approach to tackling much of the structure and bonding topic and one in which I
hope my own enthusiasm for learning ‘new stuff’ rubs off on some of my A-level chemistry
students. This approach offers the opportunity to use some of the software that is available as
free downloads; the packages ChemSketch and Jmol are used and referred to in this article.
There is little doubt that one of the most dynamic areas of scientific research at the moment is
at the interface of chemistry and biology, illustrated by the 2009 Nobel Prize for chemistry
awarded for the elucidation of the structure of the ribosome. In September 2004, with the
memories of a biochemistry degree awarded in 1986 fading fast, I decided to get up to speed
with developments by studying for an online MSc in structural biology with Birkbeck
College, University of London. Initially it was purely for my own interest, but as the course
progressed
I realised that I could address many of the chemical bonding concepts (and others) covered in
the A-level chemistry course within a protein chemistry context. In doing so, I hoped to
convey some of the excitement of cutting-edge scientific research in chemistry/biology, while
slipping in the basics under the radar. Some of this approach is considered below.
The protein folding problem
My opening pitch goes something like this:
If you want to win the next Nobel Prize for chemistry then all you have to do is solve one
problem; it’s called the protein folding problem.
All the structural information for the final 3D structure of a folded protein is contained within
its primary sequence of amino acids so, in principle, it must be possible to predict the tertiary
structure of a protein from its primary sequence. So how come no one has done it yet?
We start with a brief introduction to proteins as macromolecules made from the enzyme-
controlled polymerisation of 20 amino acids.
ChemSketch is a freely downloadable molecular imaging package from ACD Labs that has
all 20 amino acid structures pre-drawn. These can be viewed in 2D skeletal format and then
transferred to 3D. I encourage all of my students to download their own copy of this freeware
and ‘have a play’. It is excellent for getting to grips with skeletal formulae and there is a
naming function that enables students to become familiar with the IUPAC system including
the E/Z notation for geometric isomers. (Be aware that older versions use non-IUPAC
nomenclature – thus acetic, propionic, and so on, will appear if you get the wrong one.) The
most up-to-date version (12), including a Mac version, can be downloaded from the ACD
Labs site (see Websites).
The images in Figure 1, drawn using
ChemSketch, show a molecule of alanine
(2-aminopropanoic acid for the chemists) in skeletal and 3D formats. This molecule allows us
to look at the covalent bond and the π bond of the carbonyl group. There is also the option to
look at optical activity and a consideration of the fact that only one of the optical isomers
(enantiomers) is found in naturally occurring proteins.
The primary structure of a small protein may be composed of 100 amino acids, and once the
formation of the peptide bond has been looked at the main issue of the protein folding into its
active shape is considered. This gives the opportunity to look at the role of hydrogen
bonding, van der Waals interactions and ionic interactions in the stabilisation of proteins.

Figure 1 Representations of the amino acid alanine (2-aminopropanoic acid)

Consider the following. The side chain of the amino acid aspartic acid will be partially
ionised depending upon its local environment. Frequently, amino acids with acidic or basic
side groups can be found on the surface of proteins. When ionised they can form favourable
electrostatic interactions with water molecules (Figure 2).
Early attempts at determining protein structure suggested that many, if not all, acidic and
basic residues would appear on the surface of a protein where the electrostatic interactions
with water would be energetically favourable. However, consider the ionic interaction
between the aspartate group and the protonated lysine residue in Figure 3. If these amino
acids are arranged spatially close together in the final 3D structure of the protein, then such
an interaction would also be energetically favourable even within the core of the protein!
At this point, the complexity of the protein folding problem becomes clearer: the number of
possible folding permutations is astronomical.
Water aspartate
Figure 2 The electrostatic interaction of an aspartate ion with a water molecule

aspartate lysine
Figure 3 An ionic interaction between an aspartate and a lysine residue

eclipsed staggered
Figure 4 Eclipsed and staggered conformations of ethane
This gives us the opportunity to look at bond polarity. As all nuclei are surrounded by
electron clouds, the repulsive forces between these clouds would suggest that the atoms
occupy a space as far apart from each other as possible. In the Newman projections
(ChemSketch) in Figure 4, the staggered arrangement is a lower-energy spatial arrangement
than the eclipsed arrangement.
However, the polarity of particular bonds complicates the issue as an energetically favourable
dipole–dipole interaction may more than compensate for the repulsive effects.
Students might like to think about the possible stable conformations of ethane-1,2-diol. Why
might the staggered conformation not be the favoured conformation in this case?
Another important factor in determining protein structure is the hydrophobic/hydrophilic
nature of amino acid side chains. Figure 5 shows the amino acids phenylalanine and valine.
These two side groups are strongly hydrophobic and thus will tend to form induced dipole
interactions between each other and, in doing so, exclude water. It has been speculated that
this ‘clumping together’ of hydrophobic residues (the so-called
hydrophobic collapse model) is a key driving force behind the protein adopting its final 3D
structure. The significance of these interactions is illustrated when the condition of sickle-cell
anaemia is considered. A single-nucleotide mutation on the
β chain of haemoglobin leads to the replacement of a glutamic acid residue at amino acid
position 6 by a valine residue (Figure 6).
Because of this mutation, a hydrophobic patch is present on the surface of the folded

phenylalanine valine
Figure 5 Amino acids phenylalanine and valine
glutamic acid valine
Figure 6 Amino acids glutamic acid and valine
β-haemoglobin protein and the result is a clumping together of the β proteins so as to exclude
water molecules, producing the typical ‘sickling’ shape of the red blood cell (Creighton,
1997: 126).

In addition to the different bond types encountered so far, the protein folding problem also
gives the opportunity to consider the thermodynamics of protein folding. The relationship
between the total entropy change of a protein folding process can be given by the following
equation:

ΔSTOTAL = ΔSSYSTEM – ΔH/T

where ΔSTOTAL is the total entropy change, ΔSSYSTEM the entropy change associated
with the protein folding, and ΔH/T the enthalpy change associated with the folding process at
temperature T.

In order for any process to occur spontaneously, the total entropy change must be positive. In
folding from its denatured state into its active folded form, the entropy change of the system
(i.e. the protein) must be very negative (in other words, the probability of the protein adopting
its active conformation by chance alone is very small). The fact that proteins do fold
spontaneously must mean that the enthalpy change is sufficiently exothermic to compensate
for the negative entropy change of the folded protein (for details of specific experimental
evidence see Anfinsen, 1973). (Although many proteins do fold spontaneously at the
appropriate physiological conditions, many more-complicated protein structures require the
assistance of protein chaperones to achieve their folded configuration.) From this it can be
seen that the heat energy released (enthalpy change) by the formation of the specific covalent,
ionic and hydrogen bonds is a fundamental driving force in the folding of the protein into its
active conformation.
 space-filled structure

ribbon diagram

Figure 7 Two representations of a section of a human prion protein structure variant (PDB
code 1E1U)

Any folded protein then is a finely tuned structure in which covalent bonds, ionic
interactions, hydrogen bonds, van der Waals forces and hydrophobic and hydrophilic
interactions all play a vital role. The Jmol software enables the 3D visualisation of some of
these protein molecules and a greater appreciation of molecular structure. Jmol can be freely
downloaded (see Websites) and there are thousands of protein structures freely available from
the Protein Data Bank (PDB, see Websites).
Figure 7 shows two representations of a section of a human prion protein structure variant
(PDB code 1E1U). The familiar space-filled structure shows the carbon, hydrogen, oxygen,
nitrogen and sulfur atoms in grey, white, red, blue and yellow respectively. Not surprisingly,
a

protein fragment of over 12 500 atomic mass units is complex, and thus other modes of
displaying the structure are desirable. The alternative (ribbon diagram) representation of the
protein fragment, shows the secondary structures of the protein fragment in red (α-helix) and
orange (β-sheet). These various display options can be selected using the Jmol software
options.

Prions are proteins that can adopt two different forms (isoforms), a normal form and a
‘misfolded’ form. As we have seen, the number of permutations that a protein can fold into is
immense so it is not surprising that proteins can adopt more than one structure. What is
unusual about prion proteins is that, once a misfolded protein has been formed, there seems to
be a sort of domino effect whereby the other normally folded proteins convert to the
misfolded version. This can have devastating effects, including the well-documented and
fatal bovine spongiform encephalopathy (BSE) in cattle, scrapie in sheep and Creutzfeldt–
Jakob disease in humans (Prusiner, 1991). Quite how and why this happens is still an area of
intense research.

Body
 Proteins are large biomolecules and macromolecules that comprise one or more long
chains of amino acid residues. Proteins perform a vast array of functions within organisms,
including catalysing metabolic reactions, DNA replication, responding to stimuli, providing
structure to cells and organisms, and transporting molecules from one location to another.
Proteins differ from one another primarily in their sequence of amino acids, which is dictated
by the nucleotide sequence of their genes, and which usually results in protein folding into a
specific 3D structure that determines its activity.

A linear chain of amino acid residues is called a polypeptide. A protein contains at

least one long polypeptide. Short polypeptides, containing less than 20–30 residues, are rarely
considered to be proteins and are commonly called peptides. The individual amino acid
residues are bonded together by peptide bonds and adjacent amino acid residues. The
sequence of amino acid residues in a protein is defined by the sequence of a gene, which is
encoded in the genetic code. In general, the genetic code specifies 20 standard amino acids;
but in certain organisms the genetic code can include selenocysteine and—in certain archaea
—pyrrolysine. Shortly after or even during synthesis, the residues in a protein are often
chemically modified by post-translational modification, which alters the physical and
chemical properties, folding, stability, activity, and ultimately, the function of the proteins.
Some proteins have non-peptide groups attached, which can be called prosthetic groups or
cofactors. Proteins can also work together to achieve a particular function, and they often
associate to form stable protein complexes.

Once formed, proteins only exist for a certain period and are then degraded and
recycled by the cell's machinery through the process of protein turnover. A protein's lifespan
is measured in terms of its half-life and covers a wide range. They can exist for minutes or
years with an average lifespan of 1–2 days in mammalian cells. Abnormal or misfolded
proteins are degraded more rapidly either due to being targeted for destruction or due to being
unstable.

Like other biological macromolecules such as polysaccharides and nucleic acids,

proteins are essential parts of organisms and participate in virtually every process within
cells. Many proteins are enzymes that catalyse biochemical reactions and are vital to
metabolism. Proteins also have structural or mechanical functions, such as actin and myosin
in muscle and the proteins in the cytoskeleton, which form a system of scaffolding that
maintains cell shape. Other proteins are important in cell signaling, immune responses, cell
adhesion, and the cell cycle. In animals, proteins are needed in the diet to provide the
essential amino acids that cannot be synthesized. Digestion breaks the proteins down for
metabolic use.

Proteins may be purified from other cellular components using a variety of techniques
such as ultracentrifugation, precipitation, electrophoresis, and chromatography; the advent of
genetic engineering has made possible a number of methods to facilitate purification.
Methods commonly used to study protein structure and function include
immunohistochemistry, site-directed mutagenesis, X-ray crystallography, nuclear magnetic
resonance and mass spectrometry.

Proteins were recognized as a distinct class of biological molecules in the eighteenth

century by Antoine Fourcroy and others, distinguished by the molecules' ability to coagulate
or flocculate under treatments with heat or acid. Noted examples at the time included albumin
from egg whites, blood serum albumin, fibrin, and wheat gluten.

Proteins were first described by the Dutch chemist Gerardus Johannes Mulder and
named by the Swedish chemist Jöns Jacob Berzelius in 1838. Mulder carried out elemental
analysis of common proteins and found that nearly all proteins had the same empirical
formula, C400H620N100O120P1S1. He came to the erroneous conclusion that they might be
composed of a single type of (very large) molecule. The term "protein" to describe these
molecules was proposed by Mulder's associate Berzelius; protein is derived from the Greek
word πρώτειος (proteios), meaning "primary", "in the lead", or "standing in front", + -in.
Mulder went on to identify the products of protein degradation such as the amino acid leucine
for which he found a (nearly correct) molecular weight of 131 Da. Prior to "protein", other
names were used, like "albumins" or "albuminous materials" (Eiweisskörper, in German).

Early nutritional scientists such as the German Carl von Voit believed that protein
was the most important nutrient for maintaining the structure of the body, because it was
generally believed that "flesh makes flesh."[8] Karl Heinrich Ritthausen extended known
protein forms with the identification of glutamic acid. At the Connecticut Agricultural
Experiment Station a detailed review of the vegetable proteins was compiled by Thomas Burr
Osborne. Working with Lafayette Mendel and applying Liebig's law of the minimum in
feeding laboratory rats, the nutritionally essential amino acids were established. The work
was continued and communicated by William Cumming Rose. The understanding of proteins
as polypeptides came through the work of Franz Hofmeister and Hermann Emil Fischer in
1902. The central role of proteins as enzymes in living organisms was not fully appreciated
until 1926, when James B. Sumner showed that the enzyme urease was in fact a protein.

The difficulty in purifying proteins in large quantities made them very difficult for
early protein biochemists to study. Hence, early studies focused on proteins that could be
purified in large quantities, e.g., those of blood, egg white, various toxins, and
digestive/metabolic enzymes obtained from slaughterhouses. In the 1950s, the Armour Hot
Dog Co. purified 1 kg of pure bovine pancreatic ribonuclease A and made it freely available
to scientists; this gesture helped ribonuclease A become a major target for biochemical study
for the following decades.

Linus Pauling is credited with the successful prediction of regular protein secondary
structures based on hydrogen bonding, an idea first put forth by William Astbury in 1933.
Later work by Walter Kauzmann on denaturation, based partly on previous studies by Kaj
Linderstrøm-Lang, contributed an understanding of protein folding and structure mediated by
hydrophobic interactions.

The first protein to be sequenced was insulin, by Frederick Sanger, in 1949. Sanger
correctly determined the amino acid sequence of insulin, thus conclusively demonstrating that
proteins consisted of linear polymers of amino acids rather than branched chains, colloids, or
cyclols.He won the Nobel Prize for this achievement in 1958.

With the development of X-ray crystallography, it became possible to sequence

protein structures. The first protein structures to be solved were hemoglobin by Max Perutz
and myoglobin by John Kendrew, in 1958. The use of computers and increasing computing
power also supported the sequencing of complex proteins. In 1999, Roger Kornberg
succeeded in sequencing the highly complex structure of RNA polymerase using high
intensity X-rays from synchrotrons.

Since then, cryo-electron microscopy (cryo-EM) of large macromolecular assemblies

has been developed. Cryo-EM uses protein samples that are frozen rather than crystals, and
beams of electrons rather than x-rays. It causes less damage to the sample, allowing scientists
to obtain more information and analyze larger structures. Computational protein structure
prediction of small protein domains has also helped researchers to approach atomic-level
resolution of protein structures. As of 2017, the Protein Data Bank has over 126,060 atomic-
resolution structures of proteins.

Number of proteins encoded in genomes

The number of proteins encoded in a genome roughly corresponds to the number of

genes (although there may be a significant number of genes that encode RNA of protein, e.g.
ribosomal RNAs). Viruses typically encode a few to a few hundred proteins, archaea and
bacteria a few hundred to a few thousand, while eukaryotes typically encode a few thousand
up to tens of thousands of proteins (see genome size for a list of examples).

Classification

Proteins are primarily classified by sequence and structure, although other

classifications are commonly used. Especially for enzymes the EC number system provides a
functional classification scheme. Similarly, the gene ontology classifies both genes and
proteins by their biological and biochemical function, but also by their intracellular location.

Sequence similarity is used to classify proteins both in terms of evolutionary and

functional similarity. This may use either whole proteins or protein domains, especially in
multi-domain proteins. Protein domains allow protein classification by a combination of
sequence, structure and function, and thy can be combined in many different ways. In an
early study of 170,000 proteins, about two-thirds were assigned at least one domain, with
larger proteins containing more domains (e.g. proteins larger than 600 amino acids having an
average of more than 5 domains).

Biochemistry

Chemical structure of the peptide bond (bottom) and the three-dimensional structure of a
peptide bond between an alanine and an adjacent amino acid (top/inset). The bond itself is
made of the CHON elements.

Resonance structures of the peptide bond that links individual amino acids to form a protein
polymer

Most proteins consist of linear polymers built from series of up to 20 different L-α-
amino acids. All proteinogenic amino acids possess common structural features, including an
α-carbon to which an amino group, a carboxyl group, and a variable side chain are bonded.
Only proline differs from this basic structure as it contains an unusual ring to the N-end
amine group, which forces the CO–NH amide moiety into a fixed conformation. The side
chains of the standard amino acids, detailed in the list of standard amino acids, have a great
variety of chemical structures and properties; it is the combined effect of all of the amino acid
side chains in a protein that ultimately determines its three-dimensional structure and its
chemical reactivity. The amino acids in a polypeptide chain are linked by peptide bonds.
Once linked in the protein chain, an individual amino acid is called a residue, and the linked
series of carbon, nitrogen, and oxygen atoms are known as the main chain or protein
backbone.

The peptide bond has two resonance forms that contribute some double-bond
character and inhibit rotation around its axis, so that the alpha carbons are roughly coplanar.
The other two dihedral angles in the peptide bond determine the local shape assumed by the
protein backbone. The end with a free amino group is known as the N-terminus or amino
terminus, whereas the end of the protein with a free carboxyl group is known as the C-
terminus or carboxy terminus (the sequence of the protein is written from N-terminus to C-
terminus, from left to right).

The words protein, polypeptide, and peptide are a little ambiguous and can overlap in
meaning. Protein is generally used to refer to the complete biological molecule in a stable
conformation, whereas peptide is generally reserved for a short amino acid oligomers often
lacking a stable 3D structure. But the boundary between the two is not well defined and
usually lies near 20–30 residues.Polypeptide can refer to any single linear chain of amino
acids, usually regardless of length, but often implies an absence of a defined conformation.

Interactions

Proteins can interact with many types of molecules, including with other proteins,
with lipids, with carbohydrates, and with DNA.

Abundance in cells

It has been estimated that average-sized bacteria contain about 2 million proteins per
cell (e.g. E. coli and Staphylococcus aureus). Smaller bacteria, such as Mycoplasma or
spirochetes contain fewer molecules, on the order of 50,000 to 1 million. By contrast,
eukaryotic cells are larger and thus contain much more protein. For instance, yeast cells have
been estimated to contain about 50 million proteins and human cells on the order of 1 to 3
billion. The concentration of individual protein copies ranges from a few molecules per cell
up to 20 million. Not all genes coding proteins are expressed in most cells and their number
depends on, for example, cell type and external stimuli. For instance, of the 20,000 or so
proteins encoded by the human genome, only 6,000 are detected in lymphoblastoid cells.

Biosynthesis

A ribosome produces a protein using mRNA as template

The DNA sequence of a gene encodes the amino acid sequence of a protein

Proteins are assembled from amino acids using information encoded in genes. Each
protein has its own unique amino acid sequence that is specified by the nucleotide sequence
of the gene encoding this protein. The genetic code is a set of three-nucleotide sets called
codons and each three-nucleotide combination designates an amino acid, for example AUG
(adenine–uracil–guanine) is the code for methionine. Because DNA contains four
nucleotides, the total number of possible codons is 64; hence, there is some redundancy in the
genetic code, with some amino acids specified by more than one codon.[31]: 1002–42 Genes
encoded in DNA are first transcribed into pre-messenger RNA (mRNA) by proteins such as
RNA polymerase. Most organisms then process the pre-mRNA (also known as a primary
transcript) using various forms of post-transcriptional modification to form the mature
mRNA, which is then used as a template for protein synthesis by the ribosome. In
prokaryotes the mRNA may either be used as soon as it is produced, or be bound by a
ribosome after having moved away from the nucleoid. In contrast, eukaryotes make mRNA
in the cell nucleus and then translocate it across the nuclear membrane into the cytoplasm,
where protein synthesis then takes place. The rate of protein synthesis is higher in
prokaryotes than eukaryotes and can reach up to 20 amino acids per second.

The process of synthesizing a protein from an mRNA template is known as

translation. The mRNA is loaded onto the ribosome and is read three nucleotides at a time by
matching each codon to its base pairing anticodon located on a transfer RNA molecule,
which carries the amino acid corresponding to the codon it recognizes. The enzyme
aminoacyl tRNA synthetase "charges" the tRNA molecules with the correct amino acids. The
growing polypeptide is often termed the nascent chain. Proteins are always biosynthesized
from N-terminus to C-terminus.
 The size of a synthesized protein can be measured by the number of amino acids it
contains and by its total molecular mass, which is normally reported in units of daltons
(synonymous with atomic mass units), or the derivative unit kilodalton (kDa). The average
size of a protein increases from Archaea to Bacteria to Eukaryote (283, 311, 438 residues and
31, 34, 49 kDa respectively) due to a bigger number of protein domains constituting proteins
in higher organisms. For instance, yeast proteins are on average 466 amino acids long and 53
kDa in mass.The largest known proteins are the titins, a component of the muscle sarcomere,
with a molecular mass of almost 3,000 kDa and a total length of almost 27,000 amino acids.

Chemical synthesis

Short proteins can also be synthesized chemically by a family of methods known as

peptide synthesis, which rely on organic synthesis techniques such as chemical ligation to
produce peptides in high yield. Chemical synthesis allows for the introduction of non-natural
amino acids into polypeptide chains, such as attachment of fluorescent probes to amino acid
side chains. These methods are useful in laboratory biochemistry and cell biology, though
generally not for commercial applications. Chemical synthesis is inefficient for polypeptides
longer than about 300 amino acids, and the synthesized proteins may not readily assume their
native tertiary structure. Most chemical synthesis methods proceed from C-terminus to N-
terminus, opposite the biological reaction.

Structure

The crystal structure of the chaperonin, a huge protein complex. A single protein subunit is
highlighted. Chaperonins assist protein folding.

Three possible representations of the three-dimensional structure of the protein triose

phosphate isomerase. Left: All-atom representation colored by atom type. Middle: Simplified
representation illustrating the backbone conformation, colored by secondary structure. Right:
Solvent-accessible surface representation colored by residue type (acidic residues red, basic
residues blue, polar residues green, nonpolar residues white).

Most proteins fold into unique 3D structures. The shape into which a protein naturally folds is
known as its native conformation. Although many proteins can fold unassisted, simply
through the chemical properties of their amino acids, others require the aid of molecular
chaperones to fold into their native states. Biochemists often refer to four distinct aspects of a
protein's structure:
Primary structure: the amino acid sequence. A protein is a polyamide.

Secondary structure: regularly repeating local structures stabilized by hydrogen bonds. The
most common examples are the α-helix, β-sheet and turns. Because secondary structures are
local, many regions of different secondary structure can be present in the same protein
molecule.

Tertiary structure: the overall shape of a single protein molecule; the spatial relationship of
the secondary structures to one another. Tertiary structure is generally stabilized by nonlocal
interactions, most commonly the formation of a hydrophobic core, but also through salt
bridges, hydrogen bonds, disulfide bonds, and even post-translational modifications. The
term "tertiary structure" is often used as synonymous with the term fold. The tertiary
structure is what controls the basic function of the protein.

Quaternary structure: the structure formed by several protein molecules (polypeptide

chains), usually called protein subunits in this context, which function as a single protein
complex.

Quinary structure: the signatures of protein surface that organize the crowded cellular
interior. Quinary structure is dependent on transient, yet essential, macromolecular
interactions that occur inside living cells.

Proteins are not entirely rigid molecules. In addition to these levels of structure, proteins may
shift between several related structures while they perform their functions. In the context of
these functional rearrangements, these tertiary or quaternary structures are usually referred to
as "conformations", and transitions between them are called conformational changes. Such
changes are often induced by the binding of a substrate molecule to an enzyme's active site,
or the physical region of the protein that participates in chemical catalysis. In solution,
proteins also undergo variation in structure through thermal vibration and the collision with
other molecules.

Molecular surface of several proteins showing their comparative sizes. From left to right are:
immunoglobulin G (IgG, an antibody), hemoglobin, insulin (a hormone), adenylate kinase
(an enzyme), and glutamine synthetase (an enzyme).

Proteins can be informally divided into three main classes, which correlate with typical
tertiary structures: globular proteins, fibrous proteins, and membrane proteins. Almost all
globular proteins are soluble and many are enzymes. Fibrous proteins are often structural,
such as collagen, the major component of connective tissue, or keratin, the protein component
of hair and nails. Membrane proteins often serve as receptors or provide channels for polar or
charged molecules to pass through the cell membrane.

A special case of intramolecular hydrogen bonds within proteins, poorly shielded from water
attack and hence promoting their own dehydration, are called dehydrons.

Cellular functions

Proteins are the chief actors within the cell, said to be carrying out the duties specified by the
information encoded in genes. With the exception of certain types of RNA, most other
biological molecules are relatively inert elements upon which proteins act. Proteins make up
half the dry weight of an Escherichia coli cell, whereas other macromolecules such as DNA
and RNA make up only 3% and 20%, respectively.The set of proteins expressed in a
particular cell or cell type is known as its proteome.
The enzyme hexokinase is shown as a conventional ball-and-stick molecular model. To scale
in the top right-hand corner are two of its substrates, ATP and glucose.

The chief characteristic of proteins that also allows their diverse set of functions is their
ability to bind other molecules specifically and tightly. The region of the protein responsible
for binding another molecule is known as the binding site and is often a depression or
"pocket" on the molecular surface. This binding ability is mediated by the tertiary structure of
the protein, which defines the binding site pocket, and by the chemical properties of the
surrounding amino acids' side chains. Protein binding can be extraordinarily tight and
specific; for example, the ribonuclease inhibitor protein binds to human angiogenin with a
sub-femtomolar dissociation constant (<10−15 M) but does not bind at all to its amphibian
homolog onconase (>1 M). Extremely minor chemical changes such as the addition of a
single methyl group to a binding partner can sometimes suffice to nearly eliminate binding;
for example, the aminoacyl tRNA synthetase specific to the amino acid valine discriminates
against the very similar side chain of the amino acid isoleucine.

Proteins can bind to other proteins as well as to small-molecule substrates. When proteins
bind specifically to other copies of the same molecule, they can oligomerize to form fibrils;
this process occurs often in structural proteins that consist of globular monomers that self-
associate to form rigid fibers. Protein–protein interactions also regulate enzymatic activity,
control progression through the cell cycle, and allow the assembly of large protein complexes
that carry out many closely related reactions with a common biological function. Proteins can
also bind to, or even be integrated into, cell membranes. The ability of binding partners to
induce conformational changes in proteins allows the construction of enormously complex
signaling networks.: 830–49 As interactions between proteins are reversible, and depend
heavily on the availability of different groups of partner proteins to form aggregates that are
capable to carry out discrete sets of function, study of the interactions between specific
proteins is a key to understand important aspects of cellular function, and ultimately the
properties that distinguish particular cell types.

Enzymes

The best-known role of proteins in the cell is as enzymes, which catalyse chemical
reactions. Enzymes are usually highly specific and accelerate only one or a few chemical
reactions. Enzymes carry out most of the reactions involved in metabolism, as well as
manipulating DNA in processes such as DNA replication, DNA repair, and transcription.
Some enzymes act on other proteins to add or remove chemical groups in a process known as
posttranslational modification. About 4,000 reactions are known to be catalysed by enzymes.
The rate acceleration conferred by enzymatic catalysis is often enormous—as much as 1017-
fold increase in rate over the uncatalysed reaction in the case of orotate decarboxylase (78
million years without the enzyme, 18 milliseconds with the enzyme).

The molecules bound and acted upon by enzymes are called substrates. Although
enzymes can consist of hundreds of amino acids, it is usually only a small fraction of the
residues that come in contact with the substrate, and an even smaller fraction—three to four
residues on average—that are directly involved in catalysis. The region of the enzyme that
binds the substrate and contains the catalytic residues is known as the active site.

Dirigent proteins are members of a class of proteins that dictate the stereochemistry of
a compound synthesized by other enzymes.

Cellular localization

Proteins in different cellular compartments and structures tagged with green fluorescent
protein (here, white)
The study of proteins in vivo is often concerned with the synthesis and localization of the
protein within the cell. Although many intracellular proteins are synthesized in the cytoplasm
and membrane-bound or secreted proteins in the endoplasmic reticulum, the specifics of how
proteins are targeted to specific organelles or cellular structures is often unclear. A useful
technique for assessing cellular localization uses genetic engineering to express in a cell a
fusion protein or chimera consisting of the natural protein of interest linked to a "reporter"
such as green fluorescent protein (GFP).[59] The fused protein's position within the cell can
then be cleanly and efficiently visualized using microscopy,[60] as shown in the figure
opposite.

Other methods for elucidating the cellular location of proteins requires the use of known
compartmental markers for regions such as the ER, the Golgi, lysosomes or vacuoles,
mitochondria, chloroplasts, plasma membrane, etc. With the use of fluorescently tagged
versions of these markers or of antibodies to known markers, it becomes much simpler to
identify the localization of a protein of interest. For example, indirect immunofluorescence
will allow for fluorescence colocalization and demonstration of location. Fluorescent dyes are
used to label cellular compartments for a similar purpose.

Other possibilities exist, as well. For example, immunohistochemistry usually uses an

antibody to one or more proteins of interest that are conjugated to enzymes yielding either
luminescent or chromogenic signals that can be compared between samples, allowing for
localization information. Another applicable technique is cofractionation in sucrose (or other
material) gradients using isopycnic centrifugation. While this technique does not prove
colocalization of a compartment of known density and the protein of interest, it does increase
the likelihood, and is more amenable to large-scale studies.

Finally, the gold-standard method of cellular localization is immunoelectron microscopy.

This technique also uses an antibody to the protein of interest, along with classical electron
microscopy techniques. The sample is prepared for normal electron microscopic examination,
and then treated with an antibody to the protein of interest that is conjugated to an extremely
electro-dense material, usually gold. This allows for the localization of both ultrastructural
details as well as the protein of interest.

Through another genetic engineering application known as site-directed mutagenesis,

researchers can alter the protein sequence and hence its structure, cellular localization, and
susceptibility to regulation. This technique even allows the incorporation of unnatural amino
acids into proteins, using modified tRNAs, and may allow the rational design of new proteins
with novel properties.

Proteomics

The total complement of proteins present at a time in a cell or cell type is known as its
proteome, and the study of such large-scale data sets defines the field of proteomics, named
by analogy to the related field of genomics. Key experimental techniques in proteomics
include 2D electrophoresis,[66] which allows the separation of many proteins, mass
spectrometry, which allows rapid high-throughput identification of proteins and sequencing
of peptides (most often after in-gel digestion), protein microarrays, which allow the detection
of the relative levels of the various proteins present in a cell, and two-hybrid screening, which
allows the systematic exploration of protein–protein interactions. The total complement of
biologically possible such interactions is known as the interactome. A systematic attempt to
determine the structures of proteins representing every possible fold is known as structural
genomics.

Structure determination

Discovering the tertiary structure of a protein, or the quaternary structure of its complexes,
can provide important clues about how the protein performs its function and how it can be
affected, i.e. in drug design. As proteins are too small to be seen under a light microscope,
other methods have to be employed to determine their structure. Common experimental
methods include X-ray crystallography and NMR spectroscopy, both of which can produce
structural information at atomic resolution. However, NMR experiments are able to provide
information from which a subset of distances between pairs of atoms can be estimated, and
the final possible conformations for a protein are determined by solving a distance geometry
problem. Dual polarisation interferometry is a quantitative analytical method for measuring
the overall protein conformation and conformational changes due to interactions or other
stimulus. Circular dichroism is another laboratory technique for determining internal β-sheet /
α-helical composition of proteins. Cryoelectron microscopy is used to produce lower-
resolution structural information about very large protein complexes, including assembled
viruses; a variant known as electron crystallography can also produce high-resolution
information in some cases, especially for two-dimensional crystals of membrane proteins.
Solved structures are usually deposited in the Protein Data Bank (PDB), a freely available
resource from which structural data about thousands of proteins can be obtained in the form
of Cartesian coordinates for each atom in the protein.

Many more gene sequences are known than protein structures. Further, the set of solved
structures is biased toward proteins that can be easily subjected to the conditions required in
X-ray crystallography, one of the major structure determination methods. In particular,
globular proteins are comparatively easy to crystallize in preparation for X-ray
crystallography. Membrane proteins and large protein complexes, by contrast, are difficult to
crystallize and are underrepresented in the PDB. Structural genomics initiatives have
attempted to remedy these deficiencies by systematically solving representative structures of
major fold classes. Protein structure prediction methods attempt to provide a means of
generating a plausible structure for proteins whose structures have not been experimentally
determined.

Structure prediction

Constituent amino-acids can be analyzed to predict secondary, tertiary and quaternary protein
structure, in this case hemoglobin containing heme units

Complementary to the field of structural genomics, protein structure prediction develops

efficient mathematical models of proteins to computationally predict the molecular
formations in theory, instead of detecting structures with laboratory observation. The most
successful type of structure prediction, known as homology modeling, relies on the existence
of a "template" structure with sequence similarity to the protein being modeled; structural
genomics' goal is to provide sufficient representation in solved structures to model most of
those that remain. Although producing accurate models remains a challenge when only
distantly related template structures are available, it has been suggested that sequence
alignment is the bottleneck in this process, as quite accurate models can be produced if a
"perfect" sequence alignment is known. Many structure prediction methods have served to
inform the emerging field of protein engineering, in which novel protein folds have already
been designed. Also proteins (in eukaryotes ~33%) contain large unstructured but
biologically functional segments and can be classified as intrinsically disordered proteins.
Predicting and analysing protein disorder is, therefore, an important part of protein structure
characterisation.
The Shape and Structure of Proteins

From a chemical point of view, proteins are by far the most structurally complex and
functionally sophisticated molecules known. This is perhaps not surprising, once one realizes
that the structure and chemistry of each protein has been developed and fine-tuned over
billions of years of evolutionary history. We start this chapter by considering how the
location of each amino acid in the long string of amino acids that forms a protein determines
its three-dimensional shape. We will then use this understanding of protein structure at the
atomic level to describe how the precise shape of each protein molecule determines its
function in a cell.

The Shape of a Protein Is Specified by Its Amino Acid Sequence

Recall from Chapter 2 that there are 20 types of amino acids in proteins, each with different
chemical properties. A protein molecule is made from a long chain of these amino acids, each
linked to its neighbor through a covalent peptide bond (Figure 3-1). Proteins are therefore
also known as polypeptides. Each type of protein has a unique sequence of amino acids,
exactly the same from one molecule to the next. Many thousands of different proteins are
known, each with its own particular amino acid sequence.

Figure 3-1. A peptide bond.

A peptide bond. This covalent bond forms when the carbon atom from the carboxyl group of
one amino acid shares electrons with the nitrogen atom (blue) from the amino group of a
second amino acid. As indicated, a molecule of water is lost in this condensation (more...)

The repeating sequence of atoms along the core of the polypeptide chain is referred to as the
polypeptide backbone. Attached to this repetitive chain are those portions of the amino acids
that are not involved in making a peptide bond and which give each amino acid its unique
properties: the 20 different amino acid side chains (Figure 3-2). Some of these side chains are
nonpolar and hydrophobic (“water-fearing”), others are negatively or positively charged,
some are reactive, and so on. Their atomic structures are presented in Panel 3-1, and a brief
list with abbreviations is provided in Figure 3-3.

Figure 3-2. The structural components of a protein.

The structural components of a protein. A protein consists of a polypeptide backbone with

attached side chains. Each type of protein differs in its sequence and number of amino acids;
therefore, it is the sequence of the chemically different side chains (more...)
 Figure 3-3. The 20 amino acids found in proteins.

The 20 amino acids found in proteins. Both three-letter and one-letter abbreviations are listed.
As shown, there are equal numbers of polar and nonpolar side chains. For their atomic
structures, see Panel 3-1 (pp. 132–133).

As discussed in Chapter 2, atoms behave almost as if they were hard spheres with a definite
radius (their van der Waals radius). The requirement that no two atoms overlap limits greatly
the possible bond angles in a polypeptide chain (Figure 3-4). This constraint and other steric
interactions severely restrict the variety of three-dimensional arrangements of atoms (or
conformations) that are possible. Nevertheless, a long flexible chain, such as a protein, can
still fold in an enormous number of ways.

Figure 3-4. Steric limitations on the bond angles in a polypeptide chain.

Steric limitations on the bond angles in a polypeptide chain. (A) Each amino acid contributes
three bonds (red) to the backbone of the chain. The peptide bond is planar (gray shading) and
does not permit rotation. By contrast, rotation can occur about (more...)

The folding of a protein chain is, however, further constrained by many different sets of weak
noncovalent bonds that form between one part of the chain and another. These involve atoms
in the polypeptide backbone, as well as atoms in the amino acid side chains. The weak bonds
are of three types: hydrogen bonds, ionic bonds, and van der Waals attractions, as explained
in Chapter 2 (see p. 57). Individual noncovalent bonds are 30–300 times weaker than the
typical covalent bonds that create biological molecules. But many weak bonds can act in
parallel to hold two regions of a polypeptide chain tightly together. The stability of each
folded shape is therefore determined by the combined strength of large numbers of such
noncovalent bonds (Figure 3-5).

Figure 3-5. Three types of noncovalent bonds that help proteins fold.
Three types of noncovalent bonds that help proteins fold. Although a single one of these
bonds is quite weak, many of them often form together to create a strong bonding
arrangement, as in the example shown. As in the previous figure, R is used as a general
(more...)
A fourth weak force also has a central role in determining the shape of a protein. As
described in Chapter 2, hydrophobic molecules, including the nonpolar side chains of
particular amino acids, tend to be forced together in an aqueous environment in order to
minimize their disruptive effect on the hydrogen-bonded network of water molecules (see p.
58 and Panel 2-2, pp. 112–113). Therefore, an important factor governing the folding of any
protein is the distribution of its polar and nonpolar amino acids. The nonpolar (hydrophobic)
side chains in a protein—belonging to such amino acids as phenylalanine, leucine, valine, and
tryptophan—tend to cluster in the interior of the molecule (just as hydrophobic oil droplets
coalesce in water to form one large droplet). This enables them to avoid contact with the
water that surrounds them inside a cell. In contrast, polar side chains—such as those
belonging to arginine, glutamine, and histidine—tend to arrange themselves near the outside
of the molecule, where they can form hydrogen bonds with water and with other polar
molecules (Figure 3-6). When polar amino acids are buried within the protein, they are
usually hydrogen-bonded to other polar amino acids or to the polypeptide backbone (Figure
3-7).

Figure 3-6. How a protein folds into a compact conformation.

How a protein folds into a compact conformation. The polar amino acid side chains tend to
gather on the outside of the protein, where they can interact with water; the nonpolar amino
acid side chains are buried on the inside to form a tightly packed hydrophobic (more...)

Figure 3-7. Hydrogen bonds in a protein molecule.

Hydrogen bonds in a protein molecule. Large numbers of hydrogen bonds form between
adjacent regions of the folded polypeptide chain and help stabilize its three-dimensional
shape. The protein depicted is a portion of the enzyme lysozyme, and the hydrogen (more...)

Even a small protein molecule is built from thousands of atoms linked together by
precisely oriented covalent and noncovalent bonds, and it is extremely difficult to visualize
such a complicated structure without a three-dimensional display. For this reason, various
graphic and computer-based aids are used. A CD-ROM produced to accompany this book
contains computer-generated images of selected proteins, designed to be displayed and
rotated on the screen in a variety of formats.

Biologists distinguish four levels of organization in the structure of a protein. The amino acid
sequence is known as the primary structure of the protein. Stretches of polypeptide chain that
form α helices and β sheets constitute the protein’s secondary structure. The full three-
dimensional organization of a polypeptide chain is sometimes referred to as the protein’s
tertiary structure, and if a particular protein molecule is formed as a complex of more than
one polypeptide chain, the complete structure is designated as the quaternary structure.

Studies of the conformation, function, and evolution of proteins have also revealed the
central importance of a unit of organization distinct from the four just described. This is the
protein domain, a substructure produced by any part of a polypeptide chain that can fold
independently into a compact, stable structure. A domain usually contains between 40 and
350 amino acids, and it is the modular unit from which many larger proteins are constructed.
The different domains of a protein are often associated with different functions. Figure 3-12
shows an example—the Src protein kinase, which functions in signaling pathways inside
vertebrate cells (Src is pronounced “sarc”). This protein has four domains: the SH2 and SH3
domains have regulatory roles, while the two remaining domains are responsible for the
kinase catalytic activity. Later in the chapter, we shall return to this protein, in order to
explain how proteins can form molecular switches that transmit information throughout cells.
Summary

Chemical bonding is a vital topic. And it is challenging, particularly for new teachers.
Teaching bonding begins early in many curriculums and links to many later theories. It must
be addressed often and in different contexts, with appropriate pedagogical tools. This presents
a significant challenge to trainee teachers.

A recent Chemistry Education Research and Practice (CERP) article presents a literature
review on the knowledge teachers need to teach chemical bonding. Additional evidence is
presented from several Dutch teacher educators regarding their pedagogical content
knowledge (PCK) around the topic.

Knowledge of students’ understanding is particularly crucial, as they often bring

misconceptions from prior learning. Knowledge of instructional strategies (eg group learning,
concept mapping) and representations (eg models, illustrations, simulations) allows teachers
to help students develop an understanding of concepts and the relationships between them.
Appropriate representations support students in bridging macroscopic and submicroscopic
levels, and influence their ability to use structure-property relationships. Effective assessment
requires a range of formative and summative approaches to support students further their
understanding.

Living things are made up of atoms, but in most cases, those atoms aren’t just floating around
individually. Instead, they’re usually interacting with other atoms (or groups of atoms).

For instance, atoms might be connected by strong bonds and organized into molecules or
crystals. Or they might form temporary, weak bonds with other atoms that they bump into or
brush up against. Both the strong bonds that hold molecules together and the weaker bonds
that create temporary connections are essential to the chemistry of our bodies, and to the
existence of life itself.

Why form chemical bonds? The basic answer is that atoms are trying to reach the most stable
(lowest-energy) state that they can. Many atoms become stable when their valence shell is
filled with electrons or when they satisfy the octet rule (by having eight valence electrons). If
atoms don’t have this arrangement, they’ll “want” to reach it by gaining, losing, or sharing
electrons via bonds.
Ionic bonds are bonds formed between ions with opposite charges. For instance, positively
charged sodium ions and negatively charged chloride ions attract each other to make sodium
chloride, or table salt. Table salt, like many ionic compounds, doesn't consist of just one
sodium and one chloride ion; instead, it contains many ions arranged in a repeating,
predictable 3D pattern (a crystal).

Certain ions are referred to in physiology as electrolytes (including sodium, potassium, and
calcium). These ions are necessary for nerve impulse conduction, muscle contractions and
water balance. Many sports drinks and dietary supplements provide these ions to replace
those lost from the body via sweating during exercise.

Both strong and weak bonds play key roles in the chemistry of our cells and bodies. For
instance, strong covalent bonds hold together the chemical building blocks that make up a
strand of DNA. However, weaker hydrogen bonds hold together the two strands of the DNA
double helix. These weak bonds keep the DNA stable, but also allow it to be opened up for
copying and use by the cell.

More generally, bonds between ions, water molecules, and polar molecules are constantly
forming and breaking in the watery environment of a cell. In this setting, molecules of
different types can and will interact with each other via weak, charge-based attractions. For
instance, a Na ion might interact with a water molecule in one moment, and with the
negatively charged part of a protein in the next moment.

What's really amazing is to think that billions of these chemical bond interactions—strong
and weak, stable and temporary—are going on in our bodies right now, holding us together
and keeping us ticking!
Conclusion

In this article I have tried to outline an alternative context for looking at key concepts
to do with bonding, as covered in A-level and other post-16 chemistry courses such at the IB.
In addition, it has enabled the use of a number of software packages and databases that are
now freely accessible on the internet. These programs are of great benefit in helping students
to visualise what can be difficult structures. I only hope that the protein folding problem is
not solved any time soon.

This article is dedicated to William Norman Scott (1930–2011), who once bought a
rather unpromising student a book called Chemistry to sixteen by T. H. McCormack and
opened a door.

People can have strong connection , similarly some atoms can also have strong bonds
between them. ... The Proteins we need, Carbohydrates we eat are all result of chemical
bonding between atom. Gas we use in our car is a result of Chemical bonding. Oxygen ( O2 )
we breathe is a result of chemical bond.
References

https://www.ncbi.nlm.nih.gov/books/NBK26830/#:~:text=A%20protein%20molecule%20is
%20made,one%20molecule%20to%20the%20next.

Anfinsen C. B. (1973) Principles that govern the folding ofprotein chains. Science, 181, 223–
230.

Creighton, T. H. (1997) Proteins: structures and molecular properties. 2nd edn. New York:
Freeman.

Prusiner, S. B. (1991) Molecular biology of prion diseases.

Science, 252, 1515–1522.

Online MSc in structural biology, Birkbeck College, University of London:

www.cryst.bbk.ac.uk.

ChemSketch molecular imaging package: www.acdlabs. com/home.

Jmol software: jmol.sourceforge.net/download.

Protein Data Bank (PDB): www.rcsb.org/pdb/home/home.do.

https://wou.edu/chemistry/courses/online-chemistry-textbooks/ch450-and-ch451-
biochemistry-defining-life-at-the-molecular-level/chapter-2-protein-structure/

https://byjus.com/chemistry/protein-structure-and-levels-of-protein/

https://www2.chemistry.msu.edu/faculty/reusch/virttxtjml/proteins.htm

https://www.khanacademy.org/science/biology/macromolecules/proteins-and-amino-acids/a/
introduction-to-proteins-and-amino-acids