Reconstruction of a bacterial periplasmic permease in proteoliposomes and demonstration of ATP hydrolysis concomitant with transport

L. Bishop, R. Agbayani, S. V. Ambudkar, P. C. Maloney, G. Ferro-Luzzi Ames

Research output: Contribution to journalArticlepeer-review

Abstract

The histidine periplasmic permease of Salmonella typhimurium has been partially purified and reconstituted into proteoliposomes. In this in vitro preparation, transport activity is completely dependent on the presence of all four permease proteins (HisJ, HisQ, HisM, and HisP) and on internal ATP. The reconstituted system shows initial rates of transport that are comparable to those obtained with right-side-out membrane vesicles and it establishes a 100-fold concentration gradient for histidine. Proteoliposomes also transport histidine when GTP replaces ATP. Proteoliposomes do not catalyze significant ATP hydrolysis until histidine transport is initiated by addition of substrate along with HisJ, the water-soluble histidine-binding of substrate along with HisJ, the water-soluble histidine-binding protein. Both initially and throughout the course of substrate transport there is a concomitant hydrolysis of ATP, with an apparent stoichiometry (ATP/histidine) of 5:1. These experiments demonstrate directly that ATP is the source of energy for periplasmic permeases, thus resolving previous controversies on this topic.

Original languageEnglish (US)
Pages (from-to)6953-6957
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume86
Issue number18
DOIs
StatePublished - 1989

ASJC Scopus subject areas

  • General

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