Measurement of protein interaction bioenergetics: Application to structural variants of anti-sCD4 antibody

Michael L. Doyle, Michael Brigham-Burke, Michael N. Blackburn, Ian S. Brooks, Thomas M. Smith, Roland Newman, Mitchell Reff, Walter F. Stafford, Raymond W. Sweet, Alemseged Truneh, Preston Hensley, Daniel J. O'Shannessy

Research output: Contribution to journalArticle

Abstract

This chapter has described a bioenergetic analysis of the interaction of sCD4 with an IgG1, and two IgG4derivatives of an anti-sCD4 MAb. The MAbs have identical VHand VLdomains but differ markedly in their CHand CLdomains, raising the question of whether their antigen-binding chemistries are altered. We find the sCD4-binding kinetics and thermodynamics of the MAbs are indistinguishable, which indicates rigorously that the molecular details of the binding interactions are the same. We also showed the importance of using multiple biophysical methods to define the binding model before the bioenergetics can be appropriately interpreted. Analysis of the binding thermodynamics and kinetics suggests conformational changes that might be coupled to sCD4 binding by these MAbs are small or absent.

Original languageEnglish (US)
Pages (from-to)207-230
Number of pages24
JournalMethods in enzymology
Volume323
DOIs
StatePublished - Jan 1 2000
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

Cite this

Doyle, M. L., Brigham-Burke, M., Blackburn, M. N., Brooks, I. S., Smith, T. M., Newman, R., Reff, M., Stafford, W. F., Sweet, R. W., Truneh, A., Hensley, P., & O'Shannessy, D. J. (2000). Measurement of protein interaction bioenergetics: Application to structural variants of anti-sCD4 antibody. Methods in enzymology, 323, 207-230. https://doi.org/10.1016/S0076-6879(00)23368-5