Monoclonal antibody 4B1 influences the pKa of Glu325 in lactose permease (LacY) from Escherichia coli: evidence from SEIRAS

FEBS Lett. 2020 Oct;594(20):3356-3362. doi: 10.1002/1873-3468.13907. Epub 2020 Aug 30.

Abstract

The monoclonal antibody 4B1 binds to a conformational epitope on the periplasmic side of lactose permease (LacY) of Escherichia coli and inhibits H+ /lactose symport and lactose efflux under nonenergized conditions. At the same time, ligand binding and translocation reactions that do not involve net H+ translocation remain unaffected by 4B1. In this study, surface-enhanced infrared absorption spectroscopy applied to the immobilized LacY was used to study the pH-dependent changes in LacY and to access in situ the effect of the 4B1 antibody on the pKa of Glu325, the primary functional H+ -binding site in LacY. A small shift of the pK value from 10.5 to 9.5 was identified that can be corroborated with the inactivation of LacY upon 4B1 binding.

Keywords: membrane protein; monoclonal antibody 4B1; surface-enhanced infrared absorption spectroscopy.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antibodies, Monoclonal / metabolism*
  • Biological Transport
  • Escherichia coli / enzymology*
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism*
  • Glutamic Acid / metabolism*
  • Hydrogen-Ion Concentration
  • Lactose / metabolism
  • Membrane Transport Proteins / chemistry
  • Membrane Transport Proteins / metabolism*
  • Models, Molecular
  • Monosaccharide Transport Proteins / chemistry
  • Monosaccharide Transport Proteins / metabolism*
  • Spectrophotometry, Infrared
  • Symporters / chemistry
  • Symporters / metabolism*

Substances

  • Antibodies, Monoclonal
  • Escherichia coli Proteins
  • LacY protein, E coli
  • Membrane Transport Proteins
  • Monosaccharide Transport Proteins
  • Symporters
  • Glutamic Acid
  • lactose permease
  • Lactose