Role of glutamate-269 in the lactose permease of Escherichia coli.

Abstract:

:Glu-269, which is located on the hydrophilic face of putative helix VIII in the lactose permease of Escherichia coli, has been replaced with Asp, Gln or Cys by oligonucleotide-directed, site specific mutagenesis. Cells expressing Asp-269 permease exhibit no lactose accumulation or lactose-induced H+ translocation, but retain some ability to mediate lactose influx down a concentration gradient at high substrate concentrations. Furthermore, right-side-out membrane vesicles containing Asp-269 permease do not catalyse active lactose transport, facilitated lactose efflux or equilibrium exchange. Remarkably, however, Asp-269 permease accumulates beta, D-galactopyranosyl 1-thio-beta,D-galactopyranoside in a partially uncoupled fashion, whereas no transport of methyl-beta,D-thiogalactopyranoside, sucrose or maltose is detectable. Mutant permeases containing neutral replacements (Gln or Cys) or Glu-269 are completely devoid of activity, although the proteins are present in the membrane at concentrations comparable with wild-type or Asp-269 permease. The observations demonstrate that a carboxylate at position 269 is essential for transport activity, and Glu-269 is important for substrate binding and/or recognition.

journal_name

Mol Membr Biol

authors

Ujwal ML,Sahin-Tóth M,Persson B,Kaback HR

doi

10.3109/09687689409161024

subject

Has Abstract

pub_date

1994-01-01 00:00:00

pages

9-16

issue

1

eissn

0968-7688

issn

1464-5203

journal_volume

11

pub_type

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