The cyanobacterium Synechococcus sp. strain PCC 7942 contains a second alkaline phosphatase encoded by phoV

Abstract

A gene (phov) encoding an alkaline phosphatase from Synechococcus sp. strain PCC 7942 was isolated by screening a plasmid gene bank for expression of alkaline phosphatase activity in Escherichia coli JM103. Two independent clones carrying the same alkaline-phosphatase-encoding gene were isolated. One of these clones (pKW1) was further analysed and the nucleotide sequence of a contiguous 3234 bp DNA fragment was determined. Two complete open reading frames (ORFI and phoV) and an incomplete ORF3 were identified reading in the same direction. The deduced phoV gene product showed 34% identity to the alkaline phosphatase PhoA from Zymomonss mobilis, and the N-terminal part of the putative ORF3 protein exhibited 57% identity to a protein of unknown function from Frankia sp. Insertional inactivation of the Synechococcus PCC 7942 phoV gene failed, indicating an essential role for either the phoV or the ORF3 gene product. PhoV consists of 550 amino acid residues, resulting in a molecular mass of 61.3 kDa. To overexpress the Synechococcus PCC 7942 phoV gene in E. coli, plasmid pKWl was transformed into a phoA mutant of E. coli (CC118). In E. coli strain CC118(pKWI) PhoV was expressed constitutively with high rates of activity, and was shown to be membrane associated in the periplasmic space. After partial purification of the recombinant PhoV, it was shown that, like other alkaline phosphatases, the Synechococcus PhoV had a broad pH optimum in the alkaline region and a broad substrate specificity for phosphomonoesters, required Zn2+ for activity, and was inhibited by phosphate. In contrast to several other alkaline phosphatases, PhoV was inhibited by Mn2+. Due to the lack of a Synechococcus PCC 7942 phoV mutant strain, the function of PhoV remains uncertain. However, the present results show that Synechococcus PCC 7942 has a second, probably phosphate-irrepressible, alkaline phosphatase (PhoV, 61-3 kDa) in addition to the phosphate-repressible enzyme (PhoA, 145 kDa) already described.

The Chryseobacterium meningosepticum PafA enzyme: prototype of a new enzyme family of prokaryotic phosphate-irrepressible alkaline phosphatases? The GenBank accession number for pafA reported in this paper is AF157621.

“…(Wagner et al, 1995). These proteins are more closely related to each other (24n2 % sequence identity) than to any other of the ' PhoA-like ' enzymes, and are produced in a P i -irrepressible pattern (Gomez & Ingram, 1995 ;Wagner et al, 1995).…”

Ecotypic variation in phosphorus-acquisition mechanisms within marine picocyanobacteria

“…PCC 7942 (Ray et al 1991) and similar to the 149 kDa P-limitation-induced APase found in Synechocystis PCC6803 (Hirani et al 2001). Strain PCC 7942 also contains a second constitutive (noninducible) APase of 61 kDa, designated as phoV (Wagner et al 1995) for which no obvious homologues have been found within the genomes examined in this study. In contrast to WH 7803, WH 8102 encodes at least 3 APases (Palenik et al 2003): an APase (749 aa, 80 kDa) similar to sequences found in γ-proteobacteria (Shewanella and Vibrio) and 2 adjacent APase genes, a predicted APase (576 aa, 63 kDa) and a predicted APase/5'nucleotidase (750 aa, 80 kDa), both of which are homologous to the C-terminal of the WH 7803 phoA (Fig.…”

“…Furthermore, the selection of different APase genes in each strain may reflect the total range of organic substrates encountered in their respective ecological niches. In general APases and 5'-nucleotidases are capable of hydrolyzing a wide range of organic P monoesters (Wagner et al 1995, Wanner 1996. Although the substrate specificity of the APases from PCC7942 or PCC6803 have not been examined in great detail, these proteins retain domains conserved amongst alkaline phosphatases such as multiple P-loop motifs, which are thought to be involved in binding terminal PO 4 moieties and could serve to extend the range of organic P substrates hydrolyzed (Ray et al 1991, Hirani et al 2001.…”

Acclimation of unicellular cyanobacteria to macronutrient deficiency: emergence of a complex network of cellular responses

“…Furthermore, periplasmic alkaline phosphatases, for example PhoA (Ray et al, 1991) and PhoV (Wagner et al, 1995), release phosphate from various compounds, making it available for the transport systems. PhoA, an atypically large alkaline phosphatase, is strongly induced upon starvation (Ray et al, 1991).…”