Format

Send to

Choose Destination
Proc Natl Acad Sci U S A. 2008 Jul 1;105(26):8896-901. doi: 10.1073/pnas.0803577105. Epub 2008 Jun 20.

Protonation and sugar binding to LacY.

Author information

1
Department of Physiology and Microbiology, Molecular Biology Institute, University of California, Los Angeles, CA 90095-7327, USA.

Abstract

The effect of bulk-phase pH on the apparent affinity (K(d)(app)) of purified wild-type lactose permease (LacY) for sugars was studied. K(d)(app) values were determined by ligand-induced changes in the fluorescence of either of two covalently bound fluorescent reporters positioned away from the sugar-binding site. K(d)(app) for three different galactopyranosides was determined over a pH range from 5.5 to 11. A remarkably high pK(a) of approximately 10.5 was obtained for all sugars. Kinetic data for thiodigalactoside binding measured from pH 6 to 10 show that decreased affinity for sugar at alkaline pH is due specifically to increased reverse rate. A similar effect was also observed with nitrophenylgalactoside by using a direct binding assay. Because affinity for sugar remains constant from pH 5.5 to pH 9.0, it follows that LacY is fully protonated with respect to sugar binding under physiological conditions of pH. The results are consistent with the conclusion that LacY is protonated before sugar binding during lactose/H(+) symport in either direction across the membrane.

PMID:
18567672
PMCID:
PMC3021437
DOI:
10.1073/pnas.0803577105
[Indexed for MEDLINE]
Free PMC Article

Publication types, MeSH terms, Substances, Grant support

Publication types

MeSH terms

Substances

Grant support

Supplemental Content

Full text links

Icon for HighWire Icon for PubMed Central
Loading ...
Support Center