Gene cloning, expression and characterization of novel phytase from Obesumbacterium proteus.
The gene phyA encoding phytase was isolated from Obesumbacterium proteus genomic library and sequenced. The cleavage site of the PhyA signal peptide was predicted and experimentally proved. The PhyA protein shows maximum identity of 53% and 47% to phosphoanhydride phosphorylase from Yersinia pestis and phytase AppA from Escherichia coli, respectively. Based on protein sequence similarity of PhyA and its homologs, the phytases form a novel subclass of the histidine acid phosphatase family. To characterize properties of the PhyA protein, we expressed the phyA gene in E. coli. The specific activity of the purified recombinant PhyA was 310 U mg(-1) of protein. Recombinant PhyA showed activity at pH values from 1.5 through 6.5 with the optimum at 4.9. The temperature optimum was 40-45 degrees C at pH 4.9. The Km value for sodium phytate was 0.34 mM with a Vmax of 435 U mg(-1).
Zinin
Nickolay V
NV
State Research Institute of Genetics and Selection of Industrial Microorganisms, 1-st Dorozhnii proezd 1, Moscow 117545, Russian Federation. neonova1@yandex.ru
Serkina
Anna V
AV
Gelfand
Mikhail S
MS
Shevelev
Aleksei B
AB
Sineoky
Sergei P
SP
D010832:6-Phytase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification; D004755:Enterobacteriaceae / Q000201:enzymology* / Q000235:genetics; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D005798:Genes, Bacterial; D015698:Genomic Library; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013696:Temperature; D015010:Yersinia pestis / Q000201:enzymology / Q000235:genetics
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D011994:Recombinant Proteins; D010832:6-Phytase