Identification of four unique clones encoding 10 kDa proteins from Bacillus that cause phenotypic complementation of a phoA mutant strain of Escherichia coli.
A number of clones have been isolated from two Bacillus species which complement the PhoA- phenotype of Escherichia coli mutants under conditions that induce the expression of alkaline phosphatase (APase). These clones were initially thought to carry XPases because the transformed host could hydrolyse a common APase substrate, XP (5-bromo-4-chloro-3-indolyl-phosphate). The sequences of the open reading frames responsible for the phenotypic complementation showed no sequence similarity to ATPases of E. coli, human (bone-liver-kidney, intestinal or placental) or Bacillus. Therefore, these clones were designated as XPA (for X Phosphatase Activity) clones. Four of the clones encoded small (10 kDa), basic, hydrophobic proteins. Two of these, xpaB from B. subtilis 168 and xpaL2 from B. licheniformis MC14, shared 62% identity at both the DNA and the predicted amino acid sequence level. The fact that homologues from two Bacillus strains were cloned indicated that the screen was specific, but not for APase genes. It is clear that phenotypic complementation with cloned DNA from another genus does not ensure the identification of an APase gene. Possible mechanisms for the abnormal phenotypic complementation are discussed.
Lee
J W
JW
Laboratory for Molecular Biology, University of Illinois, Chicago, IL 60680.
Edwards
C W
CW
Hulett
F M
FM
D000469:Alkaline Phosphatase / Q000235:genetics* / Q000378:metabolism; D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D005798:Genes, Bacterial; D005816:Genetic Complementation Test; D006801:Humans; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010641:Phenotype; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014169:Transformation, Bacterial
D004269:DNA, Bacterial; D000469:Alkaline Phosphatase