A feasibility study of inverse contrast-matching small-angle neutron scattering method combined with size exclusion chromatography using antibody interactions as model systems

Nobuhiro Sato, Rina Yogo, Saeko Yanaka, Anne Martel, Lionel Porcar, Ken Morishima, Rintaro Inoue, Taiki Tominaga, Takao Arimori, Junichi Takagi, Masaaki Sugiyama, Koichi Kato

Research output: Contribution to journalArticlepeer-review

1 Citation (Scopus)

Abstract

Small-angle neutron scattering (SANS) and small- angle X-ray scattering (SAXS) are powerful techniques for the structural characterization of biomolecular complexes. In particular, SANS enables a selective observation of specific components in complexes by selective deuteration with contrast-matching techniques. In most cases, however, biomolecular interaction systems with heterogeneous oligomers often contain unfavorable aggregates and unbound species, hampering data interpretation. To overcome these problems, SAXS has been recently combined with size exclusion chromatography (SEC), which enables the isolation of the target complex in a multi-component system. By contrast, SEC-SANS is only at a preliminary stage. Hence, we herein perform a feasibility study of this method based on our newly developed inverse contrast-matching (iCM) SANS technique using antibody interactions as model systems. Immunoglobulin G (IgG) or its Fc fragment was mixed with 75% deuterated Fc-binding proteins, i.e. a mutated form of IgG-degrading enzyme of Streptococcus pyogenes and a soluble form of Fcγreceptor IIIb, and subjected to SEC-SANS as well as SEC-SAXS as reference. We successfully observe SANS from the non-deuterated IgG or Fc formed in complex with these binding partners, which were unobservable in terms of SANS in D2O, hence demonstrating the potential utility of the SEC-iCM-SANS approach.

Original languageEnglish
Pages (from-to)701-708
Number of pages8
JournalJournal of biochemistry
Volume169
Issue number6
DOIs
Publication statusPublished - Jun 1 2021
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

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