Investigation of Escherichia coli dimethyl sulfoxide reductase assembly and processing in strains defective for the sec-independent protein translocation system membrane targeting and translocation.
Dimethyl sulfoxide reductase is a heterotrimeric enzyme (DmsABC) localized to the cytoplasmic surface of the inner membrane. Targeting of the DmsA and DmsB catalytic subunits to the membrane requires the membrane targeting and translocation (Mtt) system. The DmsAB dimer is a member of a family of extrinsic, cytoplasmic facing membrane subunits that require Mtt in order to assemble on the membrane. We show that the MttA(2), MttB, and presumably MttA(1) but not the MttC proteins are required for targeting DmsAB to the membrane. Unlike other Mtt substrates such as trimethylamine N-oxide reductase, the soluble cytoplasmic DmsAB dimer that accumulates in the mtt deletions is very labile. Deletion of the mttA(2) or mttB genes also prevents anaerobic growth on fumarate even though fumarate reductase does not require Mtt for assembly. This was due to the lethality of membrane insertion of DmsC in the absence of the DmsAB subunits. In the absence of DmsC, DmsAB accumulates in the cytoplasm. A 45-amino acid leader on DmsA is removed during assembly. Processing does not require DmsC but does require Mtt. Translocation of DmsAB to the periplasm is not required for processing. The leader may be cleaved by a novel leader peptidase, or the long DmsA leader may traverse the membrane through the Mtt system resulting in cleavage by the periplasmic leader peptidase I followed by release of DmsA into the cytoplasm.
Sambasivarao
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D
CIHR Group in Molecular Biology of Membrane Proteins, Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Dawson
H A
HA
Zhang
G
G
Shaw
G
G
Hu
J
J
Weiner
J H
JH
D001120:Arginine / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism; D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology*; D007506:Iron-Sulfur Proteins; D010088:Oxidoreductases / Q000378:metabolism*; D010089:Oxidoreductases, N-Demethylating / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D021381:Protein Transport; D017384:Sequence Deletion
D001426:Bacterial Proteins; D017931:DNA Primers; D007506:Iron-Sulfur Proteins; D001120:Arginine; D010088:Oxidoreductases; D010089:Oxidoreductases, N-Demethylating; C023186:trimethylamine dehydrogenase; C045664:dimethyl sulfoxide reductase