The role of a purine-specific nucleoside hydrolase in spore germination of Bacillus thuringiensis.
A homologous gene (iunH) of a putative nucleoside hydrolase (NH), which had been identified from the exosporia of Bacillus cereus and Bacillus anthracis spores, was cloned from Bacillus thuringiensis subsp. kurstaki. Disruption of iunH did not affect the vegetative growth and sporulation of Bacillus thuringiensis, but promoted both inosine- and adenosine-induced spore germination. The inosine- or adenosine-induced germination rate decreased when the wild-type iunH gene was overexpressed in Bacillus thuringiensis. The iunH gene product was characterized as a purine-specific NH. The kinetic parameters of IunH with inosine as substrate were K(m)=399+/-115 microM, k(cat)=48.9+/-8.5 s(-1) and k(cat)/K(m)=1.23 x 10(5) M(-1) s(-1). The optimal pH and temperature for IunH were found to be pH 6 and 80 degrees C. Meanwhile, the specific activity of inosine hydrolase in intact spores of the wild-type strain with inosine as substrate was 2.89+/-0.23x10(-2) micromol min(-1) (mg dry wt)(-1). These results indicate that IunH is important in moderating inosine- or adenosine-induced germination of Bacillus thuringiensis spores.
Liang
Liang
L
Graduate School of Chinese Academy of Sciences, Beijing 100039, PR China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, PR China.
He
Xihong
X
Graduate School of Chinese Academy of Sciences, Beijing 100039, PR China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, PR China.
Liu
Gang
G
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, PR China.
Tan
Huarong
H
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, PR China.
D000241:Adenosine / Q000378:metabolism; D000595:Amino Acid Sequence; D001413:Bacillus thuringiensis / Q000201:enzymology* / Q000235:genetics / Q000502:physiology*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004795:Enzyme Stability; D017353:Gene Deletion; D006863:Hydrogen-Ion Concentration; D007288:Inosine / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009699:N-Glycosyl Hydrolases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013171:Spores, Bacterial / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D013696:Temperature
D004269:DNA, Bacterial; D007288:Inosine; D009699:N-Glycosyl Hydrolases; D000241:Adenosine