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PDBsum entry 3a6c
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Immune system/hydrolase
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PDB id
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3a6c
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Contents |
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107 a.a.
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114 a.a.
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129 a.a.
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References listed in PDB file
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Key reference
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Title
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Contribution of asparagine residues to the stabilization of a proteinaceous antigen-Antibody complex, Hyhel-10-Hen egg white lysozyme.
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Authors
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A.Yokota,
K.Tsumoto,
M.Shiroishi,
T.Nakanishi,
H.Kondo,
I.Kumagai.
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Ref.
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J Biol Chem, 2010,
285,
7686-7696.
[DOI no: ]
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PubMed id
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Abstract
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Many germ line antibodies have asparagine residues at specific sites to achieve
specific antigen recognition. To study the role of asparagine residues in the
stabilization of antigen-antibody complexes, we examined the interaction between
hen egg white lysozyme (HEL) and the corresponding HyHEL-10 variable domain
fragment (Fv). We introduced Ala and Asp substitutions into the Fv side chains
of l-Asn-31, l-Asn-32, and l-Asn-92, which interact directly with residues in
HEL via hydrogen bonding in the wild-type Fv-HEL complex, and we investigated
the interactions between these mutant antibodies and HEL. Isothermal titration
calorimetric analysis showed that all the mutations decreased the negative
enthalpy change and decreased the association constants of the interaction.
Structural analyses showed that the effects of the mutations on the structure of
the complex could be compensated for by conformational changes and/or by gains
in other interactions. Consequently, the contribution of two hydrogen bonds was
minor, and their abolition by mutation resulted in only a slight decrease in the
affinity of the antibody for its antigen. By comparison, the other two hydrogen
bonds buried at the interfacial area had large enthalpic advantage, despite
entropic loss that was perhaps due to stiffening of the interface by the bonds,
and were crucial to the strength of the interaction. Deletion of these strong
hydrogen bonds could not be compensated for by other structural changes. Our
results suggest that asparagine can provide the two functional groups for strong
hydrogen bond formation, and their contribution to the antigen-antibody
interaction can be attributed to their limited flexibility and accessibility at
the complex interface.
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Figure 1.
Interaction between HyHEL-10 Fv and HEL. A, overall structure
of the wild-type HyHEL-10 Fv-HEL complex. The C-α schematic
diagrams of VL, VH, and HEL are shown in green, cyan, and pink,
respectively. The residues investigated in this study are shown
in orange. Interfacial water molecules bridging Fv and HEL are
represented by red balls. B and C, local structure around the
target sites investigated in this study. Interfacial Asn
residues at sites 31, 32 (B), and 92 (C) in the VL participate
in the antigen-antibody interaction by the formation of direct
hydrogen bonds with the antigen. The contacting residues in VL
and HEL are shown by green and pink sticks, respectively. Direct
hydrogen bonds and indirect hydrogen bonds (via interfacial
water molecules) between the antigen and antibody are indicated
by red dotted lines. The figures were generated with WebLab
Viewer (Molecular Simulations Inc., San Diego).
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Figure 5.
Comparison of local structures at sites other than the
mutation site between mutant Fv-HEL and wild-type Fv-HEL
complexes. Local structures around l-Asn-92 in the LN32D-HEL
complex (A) and around l-Asn-32 in the LN92D-HEL complex (B) are
shown. Hydrogen bonds conserved in mutant and wild-type Fv-HEL
complexes are omitted to facilitate visualization. Refer to Fig.
4 for details.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2010,
285,
7686-7696)
copyright 2010.
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