A second gene for type I signal peptidase in Bradyrhizobium japonicum, sipF is located near genes involved in RNA processing and cell division

The TnphoA-induced Bradyrhizobium japonicum mutant 184 shows slow growth an d aberrant colonization of soybean nodules. Using a DNA fragment adjacent t o the transposon insertion site as a probe, a 3.4-kb Bg/II fragment of B. j aponicum 110spc4 DNA was identified and cloned. Sequence analysis indicated that two truncated ORFs and three complete ORFs were encoded on this fragm ent. A database search revealed homologies to several other prokaryotic pro teins: PdxJ (an enzyme involved in vitamin B-6 biosynthesis), AcpS (acyl ca rrier protein synthase), Lep or Sip (prokaryotic type I signal peptidase), RNase III (an endoribonuclease which processes double-stranded rRNA precurs ors and mRNA) and Era (a GTP-binding protein required for cell division). T he mutation in strain 184 was found to lie within the signal peptidase gene , which was designated sipF. Therefore, sipF is located in a region that en codes gene products involved in posttranscriptional and posttranslational p rocessing processes. By complementation of the lep(ts) E. coli mutant strai n IT41 it was demonstrated that sipF indeed encodes a functional signal pep tidase, and genetic complementation of B. japonicum mutant 184 by a 2.8-kb SalI fragment indicated that sipF; is expressed from a promoter located dir ectly upstream of sipF. Using a non-polar kanamycin resistance cassette, a specific sipF(-) mutant was constructed which exhibited defects in symbiosi s similar to those of the original mutant 184.