Overexpression of outer membrane protein OprT and increase of membrane permeability in phoU mutant of toluene-tolerant bacterium Pseudomonas putida GM730.
Eight toluene-sensitive mutants were previously isolated from the toluene-tolerant bacterium Pseudomonas putida GM730. One of these mutants was TOS6, in which Tn5 had been inserted into phoU. Susceptibility to multiple antibiotics, as well as toluene sensitivity, was increased in the phoU mutant of P. putida GM730. We compared the outer membrane proteins from the phoU mutant and wild-type via two-dimensional gel electrophoresis. A 45 kDa protein was dramatically overexpressed as the result of phoU inactivation, and this protein was identified by peptide mass fingerprinting and microsequencing as a conserved hypothetical protein consisting of 414 amino acids. The protein, designated as OprT, harbors a signal sequence and extended beta-sheets, both of which are features common to the bacterial porins. The rate of ethidium bromide accumulation in TOS6 was higher than in GM730, which indicates that the TOS6 membranes may be more permeable to ethidium bromide than are the membranes of GM730. We propose that the toluene sensitivity and increased antibiotic susceptibility observed in the phoU mutant may be attributable to increased membrane permeability.
Lee
Kyunghee
K
Division of Applied Life Science (BK21 program), Gyeongsang National University, Jinju, Republic of Korea.
Jung
Juna
J
Kim
Kwang
K
Bae
Dongwon
D
Lim
Dongbin
D
D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics*; D002463:Cell Membrane Permeability; D004251:DNA Transposable Elements; D015180:Electrophoresis, Gel, Two-Dimensional; D013058:Mass Spectrometry; D008826:Microbial Sensitivity Tests; D016254:Mutagenesis, Insertional; D011487:Protein Conformation; D021382:Protein Sorting Signals; D016958:Pseudomonas putida / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D014050:Toluene / Q000378:metabolism* / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D004251:DNA Transposable Elements; D021382:Protein Sorting Signals; D014050:Toluene