Abstract

Oligonucleotide‐directed, site‐specific mutagenesis is being applied to the problem of ion‐gradient driven active transport with the lac permease as a model system. It has been shown that Arg‐302, His‐322 and Glu‐325, neighboring residues in putative transmembrane helices IX and X, play an important role in lactose‐coupled H+ translocation, possibly as components of a catalytic triad similar to that found in the serine proteases. In addition, Cys residues, long thought to be involved in the mechanism of the permease, are not directly involved in either substrate binding or H+ translocation. Finally, a variety of mutations have no effect on permease activity indicating that single amino acid changes do not lead indiscriminately to long‐range conformational alterations.