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In and Out and Up and Down with Lac Permease

1992; Elsevier BV; Linguagem: Inglês

10.1016/s0074-7696(08)62674-1

2163-5854

H. Ronald Kaback,

Enzyme Structure and Function

This chapter highlights lactose (lac) permease transport system of Escherichia coli (E.coli). The accumulation of β-galactosides against a concentration gradient in E.coli is carried out by lac permease, which is a hydrophobic polytopic cytoplasmic membrane protein that catalyzes the coupled translocation of β-galactosides and H+ with a stoichiometry of unity. Lac permease is encoded by lac Y gene—the second structural gene in the lac operon— cloned into a recombinant plasmid, and sequenced. Sequences within the N-terminal 170 amino acid residues of lac permease are considered important for insertion. A three-amino acid sequence at the end of the last putative transmembrane helix is critical for stability and activity once the protein is inserted into the membrane. Based on the spectroscopic analyses of the purified protein and the hydropathy profiling of the amino acid sequence, a secondary structure model has been proposed in which the protein has 12 hydrophobic domains in α-helical configuration that traverse the membrane in zig-zag fashion connected by hydrophilic loops. Exclusive support for the 12-helix model has been obtained by analyzing a large number of lac permease–alkaline phosphatase (lac Y-phoA) fusions. Although the permease appears to be functional as a monomer, certain paired in-frame deletions devoid of large transmembrane domains complement functionally when expressed together in an appropriate host cell.