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Characterization of a Monomeric Heat-Labile Classical Alkaline Phosphatase from Anabaena sp. PCC7120

Ming Luo1,2, Yong-Chao Guo1, Jiao-Yu Deng1, Hong-Ping Wei1, Zhi-Ping Zhang1, Yan Leng1,2, Dong Men1,2, Li-Rong Song3, Xian-En Zhang1, and Ya-Feng Zhou1*

1State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China; fax: +86(027)8719-9492; E-mail: zyf@wh.iov.cn; luoming1976@yahoo.cn; guoyc@ust.hk; dengjy@wh.iov.cn; hpwei@wh.iov.cn; zhangzp@wh.iov.cn; x.zhang@wh.iov.cn

2Graduate School, Chinese Academy of Science, Beijing 100039, China; audreyyanleng@yahoo.com.cn; Winter_J_Gates@yahoo.com.cn

3State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, 430072, China; lrsong@ihb.ac.cn

* To whom correspondence should be addressed.

Received March 12, 2010; Revision received March 30, 2010
Alkaline phosphatases (APs), known inducible enzymes of the Pho regulon and poorly characterized in cyanobacteria, hydrolyze phosphomonoesters to produce inorganic phosphate (Pi) during Pi starvation. In this study, two predicted alkaline phosphatase genes in the genome of Anabaena sp. PCC 7120, all2843 and alr5291, were apparently induced during Pi starvation. Sequence analysis showed that alr5291 encodes a protein that is an atypical alkaline phosphatase like other cyanobacteria PhoAs, but the protein encoded by all2843 is very similar to the classical PhoAs, such as Escherichia coli alkaline phosphatase (EAP). To date, there have been no reports about classical phoA in cyanobacterial genomes. The alkaline phosphatase APA, coded by all2843, is characterized as a metalloenzyme containing Mg2+ and Zn2+ with molar ratio of 1 : 2. Site-directed mutagenesis analysis indicated that, though the active center of APA is highly conserved in comparison with EAP, differences do exist between APA and EAP in metal ion coordination. Besides, biochemical analysis revealed that APA is a monomeric protein and inactivated rapidly at 50°C. These results suggest that APA is the first monomeric heat-labile classical PhoA found in cyanobacteria.
KEY WORDS: Anabaena sp. PCC7120, phosphorous starvation, alkaline phosphatase, metalloenzyme, site-directed mutagenesis

DOI: 10.1134/S0006297910050172