Abstract
The major facilitator superfamily (MFS) is a diverse group of secondary transporters with members found in all kingdoms of life. A paradigm for MFS is the lactose permease (LacY) ofEscherichia coli, which couples the stoichiometric translocation of a galactopyranoside and anH+across the cytoplasmic membrane. LacY has been the test bed for the development of many methods applied for the analysis of transport proteins. X-ray structures of an inward-facing conformation and the most recent structure of an almost occluded conformation confirm many conclusions from previous studies. Although structure models are critical, they are insufficient to explain the catalysis of transport. The clues to understanding transport are based on the principles of enzyme kinetics. Secondary transport is a dynamic process—static snapshots of X-ray crystallography describe it only partially. However, without structural information, the underlying chemistry is virtually impossible to conclude. A large body of biochemical/biophysical data derived from systematic studies of site-directed mutants in LacY suggests residues critically involved in the catalysis, and a working model for the symport mechanism that involves alternating access of the binding site is presented. The general concepts derived from the bacterial LacY are examined for their relevance to other MFS transporters.
Highlights
Proteins can act as molecular devices to convert one energy form to another through cycles of conformational transitions
In addition to regulatory loci, it is widely known that the lac operon contains three structural genes: (i) the Z gene encoding galactosidase, a cytosolic enzyme that catalyzes cleavage of lactose into galactose and glucose once it enters the cell; (ii) the Y gene encoding LacY; and (iii) the A gene encoding galactoside transacetylase, a cytosolic enzyme that catalyzes acetylation of mainly thiogalactopyranosides with acetyl-CoA as the acetyl donor and has an unknown physiological function
Lactose permease belongs to the major facilitator superfamily (MFS; Transporter Classification Database, TCDB #2.A.1), the largest family of secondary transporters, present in all kingdoms of life
Summary
Proteins can act as molecular devices to convert one energy form to another through cycles of conformational transitions. Symporters, antiporters, transporters, carriers, or permeases are such molecular devices that catalyze substrate-specific equilibration and/or translocation of solutes across a biological membrane (Figure 1(a)). In 1955, when the existence of cell membranes themselves was still being questioned, let alone the existence of proteins that transport solutes across them, Cohen and Rickenberg [1] found an inducible transport system for lactose in Escherichia coli, and it was subsequently found to be part of the famous lac operon [2]. (b) Uphill substrate (S) accumulation in response to an inwardly in response to ΔμH+ generated by either H+ directed downhill lactose gradient (influx). Downhill sugar influx with the generation of or efflux will drive ΔμH+ , the polarity of which depends on the direction of the lactose gradient (Figures 1(c) and 1(d)). −60 mV in the form of either a Δψ, a ΔpH, or a combination of both will theoretically generate a tenfold concentration
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