A 25-kDa beta-lactam-induced outer membrane protein of Vibrio cholerae. Purification and characterization.
A 25-kDa outer membrane protein, induced following treatment of Vibrio cholerae cells with beta-lactam antibiotics and constituting about 8-10% of the total outer membrane proteins of beta-lactam-resistant mutants, has been purified to homogeneity. It is a basic (pI 8.5) protein rich in beta-sheet structure and is a homodimer, the monomers being held together by hydrophobic interactions. The effective hydrophobicity of the protein is low, and a large part of the protein is exposed on the surface of the outer membrane. The protein does not have beta-lactamase or autolytic activity and is not a penicillin-binding protein. The Stoke's radius of the 25-kDa protein (26 A) is comparable to the pore size of the V. cholerae OmpF-like porin. Proteoliposome swelling assay showed that the 25-kDa protein might block the pores of OmpF through which beta-lactam antibiotics normally enter the cells. Twenty-two amino acid residues from the N-terminal end of the 25-kDa protein have been sequenced, and a 32-mer oligonucleotide probe was synthesized using the amino acid residues 2-12. This probe was used to identify the gene encoding the 25-kDa protein. The beta-lactam-resistant cells are insensitive to changes in the osmolarity of the growth medium in contrast to the wild type cells which exhibit osmoregulation of OmpF and OmpC synthesis. All beta-lactam-resistant mutants examined are resistant to novobiocin.
Deb
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Department of Biophysics, Indian Institute of Chemical Biology, Calcutta.
Bhattacharyya
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Das
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D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D002458:Cell Fractionation / Q000379:methods; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004591:Electrophoresis, Polyacrylamide Gel; D005798:Genes, Bacterial; D008081:Liposomes; D008970:Molecular Weight; D009675:Novobiocin / Q000494:pharmacology; D015345:Oligonucleotide Probes; D010403:Penicillin Resistance; D011510:Proteolipids / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry; D014734:Vibrio cholerae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D047090:beta-Lactams
D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D008081:Liposomes; D015345:Oligonucleotide Probes; D011510:Proteolipids; D047090:beta-Lactams; C020485:proteoliposomes; D009675:Novobiocin