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Pilani Campus

CHEM F343: Inorganic Chemistry III

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Bioinorganic Chemistry
Structure similarity of Hb and Mb

Tertiary structure of a and b are remarkably similar

both to each other and to Mb

Only 18% of the corresponding residues are identical

among these 3 polypeptides
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Structure similarity of Hb and Mb

The 3-D structures of the a and b chains of Hb look like

that of Mb,
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but their primary sequences are different.
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Hemerythrin (Hr) and Hemocyanine (Hc)
•are found in invertebrates (Annelids, Mollusks, Arthropods)
•difficult to crystallize (single crystal) as it aggregates
• Hr isolated from the blood of invertebrates is typically in
form of octamer
• Hc exists as large aggregates
•Chemical and physical properties of Hr and Hc are available.

Terminology for the oxidation states of metal ions

Hemerythrin
Fe(II).Fe(II) (deoxy)
Fe(III).Fe(III) (met)
Fe(II).Fe(III) (Semi-met)
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Hemocyanine
Cu(I).Cu(I) (deoxy)
Cu(II).Cu(II) (met)
Cu(I).Cu(II)(Semi-met)
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Hemerythrin (Hr) and Hemocyanine (Hc)

• O2 binding is essentially non cooperative for Hr

• Cooperative binding of O2 is common for the multimeric Hc
• In both cases, O2 binding is accompanied by two electron
oxidation of the binuclear center to produce bound peroxide
• Raman spectroscopy can be used to detect the nature of
O2 binding

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Hemerythrin
O2  Fe(III) results in a peak at 844 cm-1 indicating
peroxide type binding

• Binding mode was

explored using
asymmetrically labeled
Fe—16O—18O and
doublet was obtained

Resonance Raman spectrum of Fe—16O—18O; central peak can

be fitted with two smooth curves for Fe—16O—18O and Fe—
18O—16O
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Hemerythrin

Possible binding mode

•III and IV: considered due to unsymmetrical mode

• IV is ultimately considered based on single crystal X-ray
diffraction along with other routine studies
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O2 binding to Hemerythrin

1.High spin Fe(II) ions 2. Antiferromagnetically coupled

No proton release and hence no Bohr effect

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 Hemocyanin: Structure

Reversible oxygen binding derives from two features of the

binding pocket.

1. 2 Cu+ ions are each bound by 3 His residues.

Geometry: pseudo Td. With His forming base, Cu at the
Apex, favoring Cu+ not Cu2+( favors Sq. pl. geometry)

2. Cavity lined with

hydrophobic residues
disfavor charged int..
Pocket size reduces
from 0.95 to 0.72 Å,
making 2 Cu ions
closer by 0.2 - 0.3 Å.

Blue
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Enzymes: 6 types
• Oxidoreductase: Oxidation/ reduction
• Transferase: Transfers the group from one compound to
another
•Hydrolase: hydrolysis
• Isomerase: converts substrate into it’s isomeric form
• Lyase: non hydrolytic addition or removal of group
• Ligase: Synthesis of a large molecules from two smaller ones
Zn—OH2  Zn—OH- + H+ pKa = 8.8
Mg—OH2  Mg—OH- + H+ pKa = 11.4

Zinc is stronger Lewis acid

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Carboxypeptidase A
• General Rule
• Zn(II) is employed where substrate bear a carbonyl
(C=O) functional group (ester, amide etc.)
• Mg(II) acts as cofactor for the enzymes responsible
for hydrolysis or formation of phosphate esters

Carboxypeptidase A
• Catalyzes the hydrolysis of C-terminal amino acid of any
polypeptide and prefers for substrate with aromatic side
chain
• Active site possesses hydrophobic pocket to recognize
such side chain
• Bovine Carboxypeptidase A has been extensively
explored
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Carboxypeptidase A

• Amide carbonyl is hydrogen bonded to Arg

• Attack of H2O is assisted by glutamate and Zn(II)
• Zn(II) does not interact directly with the C=O oxygen.
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Carboxypeptidase A

Tetrahedral intermediate is stabilized and then collapse

with proton abstraction by Glu
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Carboxypeptidase A

Electrostatic interactions between the product

carboxylate and Glu 270 assists product release
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From: Bertini, I.; Gray, H.B.; Lippard, S.J.; Valentine, J.S.: "Bioinorganic
Chemistry" (1994), University Science Books, pg. 83

(Chemistry Library, QP531 .B543 1994 )

Alkaline Phosphatase

• Zn(II)---Zn(II) = 4 Å
• One Mg (II) ion is
located at 5-7 Å away
from both the Zn(II)
ion
• Zn(II) ions are bound
with His and Asp
• One Zn(II) is close to
Ser-OH, which acts as
nucleophile to attack
the phosphate ester
• TBP geometry about
Zn(II)
Dimer of 94 kDa
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Alkaline Phosphatase

J. Mol. Biol. 2012, 415, p102-17

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Alkaline Phosphatase

PROTEIN SC. 2010, 19, p75—84

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Alkaline Phosphatase

• Optimum activity at pH 8
• One Zn(II) is closely
associated with Ser while
other is with water
• Water coordinated Zn(II)
stabilizes the -ve charge
developed on alkoxide group
produced
• The pKa of aquated
Zn(II) is 8.8, which yields
R-OH and displaces the
phosphoryl group from the
phosphoserin intermediate
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Alkaline Phosphatase

• K3 is the
slowest step
• ROH is rapidly
produced
• Burst kinetics:
Due to an initial
high velocity of
enzymatic
turnover when
adding enzyme
to substrate.
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