Lac permease of Escherichia coli: on the path of the proton.
Lactose/H+ symport in Escherichia coli is catalysed by a hydrophobic transmembrane protein encoded by the lacY gene that has been purified to homogeneity, reconstituted into proteoliposomes and shown to be completely functional as a monomer. Circular dichroic studies and hydropathy profiling of the amino-acid sequence of this 'lac' permease suggest a secondary structure in which the polypeptide consists of 12 hydrophobic segments in alpha-helical conformation that traverse the membrane in zig-zag fashion connected by shorter, hydrophilic domains with most of the charged residues and many of the residues commonly found in beta-turns. Support for certain general aspects of the model has been obtained from other biophysical studies, as well as biochemical, immunological and genetic approaches. Oligonucleotide-directed, site-specific mutagenesis is currently being utilized to probe the structure and function of the permease. Application of the technique provides an indication that Arg302 (putative helix IX), His322 (putative helix X) and Glu325 (putative helix X) may be sufficiently close to hydrogen-bond and that these residues play a critical role in lactose-coupled H+ translocation, possibly as components of a charge-relay type of mechanism. In contrast, Cys residues, which were long thought to play a central role in the mechanism of lactose/H+ symport, do not appear to be involved in either substrate binding or H+ translocation.
Kaback
H R
HR
Department of Biochemistry, Roche Research Center, Nutley, New Jersey 07110.
D000595:Amino Acid Sequence; D002462:Cell Membrane / Q000201:enzymology; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D011487:Protein Conformation; D011522:Protons; D027981:Symporters
D029968:Escherichia coli Proteins; C050221:LacY protein, E coli; D026901:Membrane Transport Proteins; D009004:Monosaccharide Transport Proteins; D011522:Protons; D027981:Symporters; C021499:lactose permease