Organization of dimethyl sulfoxide reductase in the plasma membrane of Escherichia coli.
Dimethyl sulfoxide reductase is a trimeric, membrane-bound, iron-sulfur molybdoenzyme induced in Escherichia coli under anaerobic growth conditions. The enzyme catalyzes the reduction of dimethyl sulfoxide, trimethylamine N-oxide, and a variety of S- and N-oxide compounds. The topology of dimethyl sulfoxide reductase subunits was probed by a combination of techniques. Immunoblot analysis of the periplasmic proteins from the osmotic shock and chloroform wash fluids indicated that the subunits were not free in the periplasm. The reductase was susceptible to proteases in everted membrane vesicles, but the enzyme in outer membrane-permeabilized cells became protease sensitive only after detergent solubilization of the E. coli plasma membrane. Lactoperoxidase catalyzed the iodination of each of the three subunits in an everted membrane vesicle preparation. Antibodies to dimethyl sulfoxide reductase and fumarate reductase specifically agglutinated the everted membrane vesicles. No TnphoA fusions could be found in the dmsA or -B genes, indicating that these subunits were not translocated to the periplasm. Immunogold electron microscopy of everted membrane vesicles and thin sections by using antibodies to the DmsABC, DmsA, DmsB subunits resulted in specific labeling of the cytoplasmic surface of the inner membrane. These results show that the DmsA (catalytic subunit) and DmsB (electron transfer subunit) are membrane-extrinsic subunits facing the cytoplasmic side of the plasma membrane.
Sambasivarao
D
D
Department of Biochemistry, University of Alberta, Edmonton, Canada.
Scraba
D G
DG
Trieber
C
C
Weiner
J H
JH
D000906:Antibodies; D002462:Cell Membrane / Q000201:enzymology* / Q000648:ultrastructure; D004251:DNA Transposable Elements; D004926:Escherichia coli / Q000201:enzymology* / Q000648:ultrastructure; D015151:Immunoblotting; D007506:Iron-Sulfur Proteins; D007700:Kinetics; D008854:Microscopy, Electron; D010088:Oxidoreductases / Q000032:analysis / Q000378:metabolism*
D000906:Antibodies; D004251:DNA Transposable Elements; D007506:Iron-Sulfur Proteins; D010088:Oxidoreductases; C045664:dimethyl sulfoxide reductase