Cloning, sequencing and characterization of a novel phosphatase gene, phoI, from soil bacterium Enterobacter sp. 4.
A gene, phoI, coding for a phosphatase from Enterobacter sp. 4 was cloned in Escherichia coli and sequenced. Analysis of the sequence revealed one open reading frame (ORF) that encodes a 269-amino acid protein with a calculated molecular mass of 29 kDa. PhoI belongs to family B acid phosphatase and exhibits 49.4% identity and 62.4% homology to the hel gene from Heamophilus influenzae, which encoded an outer membrane protein (P4). The optimum pH and temperature for phosphatase activity were pH 5.5 and 40 degrees C, respectively. Its specific activity on rho-nitrophenyl phosphatate was 70 U/mg at pH 5.5 and 40 degrees C. Enzyme activity was inhibited by Al3+, EDTA, and DTT, but fivefold activated by Cu2+ ion (350 U/mg). PhoI showed a strong synergistic effect when used with a purified E. coli phytase, AppA, to estimate combination effects.
Kang
Seung Ha
SH
School of Agricultural Biotechnology, Seoul National University Seoul, 151-921, Korea.
Cho
Kwang Keun
KK
Bok
Jin Duck
JD
Kim
Sung Chan
SC
Cho
Jaie Soon
JS
Lee
Peter Chang-Whan
PC
Kang
Sang Kee
SK
Lee
Hong Gu
HG
Woo
Jung Hee
JH
Lee
Hyun Jeong
HJ
Lee
Sang Cheol
SC
Choi
Yun Jaie
YJ
D010832:6-Phytase / Q000378:metabolism; D000135:Acid Phosphatase / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D003001:Cloning, Molecular; D004754:Enterobacter / Q000201:enzymology*; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000378:metabolism; D004950:Esterases / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008074:Lipoproteins / Q000235:genetics; D008969:Molecular Sequence Data; D010710:Phosphates / Q000378:metabolism; D010833:Phytic Acid / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity; D013696:Temperature
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; D008074:Lipoproteins; D010710:Phosphates; C098614:e(P4) lipoprotein, Haemophilus influenzae; D010833:Phytic Acid; D004950:Esterases; C070989:phosphoenolpyruvate phosphatase; D000135:Acid Phosphatase; D010832:6-Phytase; C406788:appA protein, E coli