Cloning of the Bacillus firmus OF4 cls gene and characterization of its gene product.
The gene that codes for cardiolipin (CL) synthase and an adjacent gene that codes for a MecA homolog in the alkaliphilic bacteria Bacillus firmus OF4 have been cloned and sequenced (GenBank accession number U88888). The cls gene contains 1509 nucleotides, corresponding to a polypeptide of 57.9 kDa. The predicted amino acid sequence has 129 identities and 100 similarities with the Escherichia coli CL synthase. Homologies were also noted with polypeptide sequences from putative cls genes from Bacillus subtilis and Pseudomonas putida. Conserved histidine, tyrosine, and serine residues may be part of the active site and participate in phosphatidyl group transfer. The B. firmus OF4 cls gene product was inserted into plasmid pET3 to form a recombinant plasmid pDG2, which overproduces CL synthase in E. coli. A membrane fraction containing the overproduced enzyme converts phosphatidylglycerol to CL and glycerol. The B. firmus enzyme is stimulated by potassium phosphate, inhibited by CL and phosphatidate, and has a slightly higher pH optimum than the E. coli enzyme.
Guo
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Queens College CUNY, Department of Chemistry and Biochemistry, Flushing, NY 11367, USA.
Tropp
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BE
D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D002308:Cardiolipins / Q000096:biosynthesis; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D006863:Hydrogen-Ion Concentration; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D010710:Phosphates / Q000494:pharmacology; D010712:Phosphatidic Acids / Q000032:analysis / Q000494:pharmacology; D010715:Phosphatidylglycerols / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D017855:Transferases (Other Substituted Phosphate Groups) / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D002308:Cardiolipins; D008565:Membrane Proteins; D010710:Phosphates; D010712:Phosphatidic Acids; D010715:Phosphatidylglycerols; D011994:Recombinant Proteins; D017855:Transferases (Other Substituted Phosphate Groups); C100091:cardiolipin synthetase