Contribution of asparagine residues to the stabilization of a proteinaceous antigen-antibody complex, HyHEL-10-Hen egg white lysozyme

Akiko Yokota, Kouhei Tsumoto, Mitsunori Shiroishi, Takeshi Nakanishi, Hidemasa Kondo, Izumi Kumagai

研究成果: ジャーナルへの寄稿記事

13 引用 (Scopus)

抄録

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.

元の言語英語
ページ(範囲)7686-7696
ページ数11
ジャーナルJournal of Biological Chemistry
285
発行部数10
DOI
出版物ステータス出版済み - 3 5 2010

Fingerprint

Egg White
Asparagine
Muramidase
Antigen-Antibody Complex
Hydrogen
Hydrogen bonds
Stabilization
Antigens
Antibodies
Mutation
Immunoglobulin Variable Region
Antibody Affinity
Hydrogen Bonding
Viperidae
Germ Cells
Titration
Functional groups
Enthalpy
Substitution reactions
hen egg lysozyme

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

これを引用

Contribution of asparagine residues to the stabilization of a proteinaceous antigen-antibody complex, HyHEL-10-Hen egg white lysozyme. / Yokota, Akiko; Tsumoto, Kouhei; Shiroishi, Mitsunori; Nakanishi, Takeshi; Kondo, Hidemasa; Kumagai, Izumi.

:: Journal of Biological Chemistry, 巻 285, 番号 10, 05.03.2010, p. 7686-7696.

研究成果: ジャーナルへの寄稿記事

Yokota, Akiko ; Tsumoto, Kouhei ; Shiroishi, Mitsunori ; Nakanishi, Takeshi ; Kondo, Hidemasa ; Kumagai, Izumi. / Contribution of asparagine residues to the stabilization of a proteinaceous antigen-antibody complex, HyHEL-10-Hen egg white lysozyme. :: Journal of Biological Chemistry. 2010 ; 巻 285, 番号 10. pp. 7686-7696.
@article{1960cbcfe82a458f8482605e124b235c,
title = "Contribution of asparagine residues to the stabilization of a proteinaceous antigen-antibody complex, HyHEL-10-Hen egg white lysozyme",
abstract = "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.",
author = "Akiko Yokota and Kouhei Tsumoto and Mitsunori Shiroishi and Takeshi Nakanishi and Hidemasa Kondo and Izumi Kumagai",
year = "2010",
month = "3",
day = "5",
doi = "10.1074/jbc.M109.089623",
language = "English",
volume = "285",
pages = "7686--7696",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "10",

}

TY - JOUR

T1 - Contribution of asparagine residues to the stabilization of a proteinaceous antigen-antibody complex, HyHEL-10-Hen egg white lysozyme

AU - Yokota, Akiko

AU - Tsumoto, Kouhei

AU - Shiroishi, Mitsunori

AU - Nakanishi, Takeshi

AU - Kondo, Hidemasa

AU - Kumagai, Izumi

PY - 2010/3/5

Y1 - 2010/3/5

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=77951215193&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=77951215193&partnerID=8YFLogxK

U2 - 10.1074/jbc.M109.089623

DO - 10.1074/jbc.M109.089623

M3 - Article

C2 - 20038580

AN - SCOPUS:77951215193

VL - 285

SP - 7686

EP - 7696

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 10

ER -