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PDBsum entry 1g7h
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Hydrolase inhibitor/hydrolase
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PDB id
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1g7h
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Contents |
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107 a.a.
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116 a.a.
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129 a.a.
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Estimation of the hydrophobic effect in an antigen-Antibody protein-Protein interface.
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Authors
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E.J.Sundberg,
M.Urrutia,
B.C.Braden,
J.Isern,
D.Tsuchiya,
B.A.Fields,
E.L.Malchiodi,
J.Tormo,
F.P.Schwarz,
R.A.Mariuzza.
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Ref.
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Biochemistry, 2000,
39,
15375-15387.
[DOI no: ]
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PubMed id
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Abstract
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Antigen-antibody complexes provide useful models for analyzing the
thermodynamics of protein-protein association reactions. We have employed
site-directed mutagenesis, X-ray crystallography, and isothermal titration
calorimetry to investigate the role of hydrophobic interactions in stabilizing
the complex between the Fv fragment of the anti-hen egg white lysozyme (HEL)
antibody D1.3 and HEL. Crystal structures of six FvD1.3-HEL mutant complexes in
which an interface tryptophan residue (V(L)W92) has been replaced by residues
with smaller side chains (alanine, serine, valine, aspartate, histidine, and
phenylalanine) were determined to resolutions between 1.75 and 2.00 A. In the
wild-type complex, V(L)W92 occupies a large hydrophobic pocket on the surface of
HEL and constitutes an energetic "hot spot" for antigen binding. The losses in
apolar buried surface area in the mutant complexes, relative to wild-type, range
from 25 (V(L)F92) to 115 A(2) (V(L)A92), with no significant shifts in the
positions of protein atoms at the mutation site for any of the complexes except
V(L)A92, where there is a peptide flip. The affinities of the mutant Fv
fragments for HEL are 10-100-fold lower than that of the original antibody.
Formation of all six mutant complexes is marked by a decrease in binding
enthalpy that exceeds the decrease in binding free energy, such that the loss in
enthalpy is partly offset by a compensating gain in entropy. No correlation was
observed between decreases in apolar, polar, or aggregate (sum of the apolar and
polar) buried surface area in the V(L)92 mutant series and changes in the
enthalpy of formation. Conversely, there exist linear correlations between
losses of apolar buried surface and decreases in binding free energy (R(2) =
0.937) as well as increases in the solvent portion of the entropy of binding
(R(2) = 0.909). The correlation between binding free energy and apolar buried
surface area corresponds to 21 cal mol(-1) A(-2) (1 cal = 4.185 J) for the
effective hydrophobicity at the V(L)92 mutation site. Furthermore, the slope of
the line defined by the correlation between changes in binding free energy and
solvent entropy approaches unity, demonstrating that the exclusion of solvent
from the binding interface is the predominant energetic factor in the formation
of this protein complex. Our estimate of the hydrophobic contribution to binding
at site V(L)92 in the D1.3-HEL interface is consistent with values for the
hydrophobic effect derived from classical hydrocarbon solubility models. We also
show how residue V(L)W92 can contribute significantly less to stabilization when
buried in a more polar pocket, illustrating the dependence of the hydrophobic
effect on local environment at different sites in a protein-protein interface.
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