Overproduction and identification of the ftsQ gene product, an essential cell division protein in Escherichia coli K-12.

摘要

ftsQ is an essential cell division gene in Escherichia coli. The ftsQ gene has been sequenced, and a presumptive open reading frame has been identified; however, no protein product has been observed (A.C. Robinson, D.J. Kenan, G.F. Hatfull, N.F. Sullivan, R. Spiegelberg, and W.D. Donachie, J. Bacteriol. 160:546-555, 1984, and Q.M. Yi, S. Rockenbach, J.E. Ward, Jr., and J. Lutkenhaus, J. Mol. Biol. 184:399-412, 1985). The ftsQ gene was isolated on a 970-base-pair EcoRI-PvuII fragment of the E. coli chromosome and used to construct a trp-lac (Ptac) transcriptional fusion in plasmid pKK223-3. The fused construct (pDSC78) complemented an ftsQ1(Ts) mutant strain in trans, restoring growth at 42 degrees C on low-salt medium. An ftsQ1(Ts) mutant strain transformed with pDSC78 appeared normal upon microscopic examination, with no indication of filamentation. The ftsQ gene product was identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and two-dimensional gel electrophoresis of radiolabeled, isopropyl-beta-D-thiogalactopyranoside-induced maxicell and normal cell extracts. As predicted from the nucleotide sequence, the 970-base-pair EcoRI-PvuII fragment encoded a polypeptide of approximately 31,400 daltons. Analysis of the data obtained from pulse-chase experiments in maxicells and normal cells suggests that the FtsQ protein is stable. Most of the radiolabeled FtsQ protein from maxicells was found in the inner membrane. On the basis of available information, the prior inability to detect FtsQ can be attributed to low levels of transcription or translation rather than to proteolysis.