Phosphate-irrepressible Alkaline Phosphatase of Zymomonas mobilis

Abstract

The phosphate regulation and subcellular location of the hydrolytic enzyme alkaline phosphatase were investigated in the Gram-negative bacterium Zymomoms mobilk. The biosynthesis of alkaline phosphatase was not derepressed at a low phosphate concentration as is generally observed in other micro-organisms, nor was it repressed by high phosphate concentrations in the medium. The enzyme level was rather constant during the growth phases in batch culture, at a value at least 8-4-fold lower than that observed in Escherichia coli. The alkaline phosphatase of Z. mobilis was found associated with the membrane fraction after cell disruption, osmotic shock treatment or spheroplast formation. This is a rather unusual location, since most of the alkaline phosphatases from Gram-negative bacteria have been shown to be periplasmic enzymes. Activity staining on polyacrylamide gels after two-dimensional electrophoresis revealed two isoforms of alkaline phosphatase, each of approximate molecular mass 56 kDa. These two forms belong to a group including the more basic envelope proteins of 2. mobilis. Our results indicate that the enzyme from 2. mobiZis differs markedly from typical alkaline phosphatases of other Gram-negative bacteria.

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

“…The APase of Antarctic bacteria was inactivated by incubation for 10 min at 55 "C (Kobori e t al., 1984). The PhoA of 2. mobilir ZM1 remained stable for 1 h at 60 O C , but further increase in temperature caused inactivation (Michel & Baratti, 1989).…”

“…However, from the information available in the literature, it is not clear whether the APase from 2. mobilis CP4 (database accession number L36230; P. F. Gomez & L. 0. Ingram, unpublished, 1994), which shows 34% identity to the Synechococcus PCC 7942 PhoV, is identical with the APase isolated and characterized from 2. mobilis ZM1 (Michel & Baratti, 1989;Michel et al, 1992;Baoudene-Assali et al, 1993).…”

Purification and properties of a phosphate-irrepressible membrane-bound alkaline phosphatase from Zymomonas mobilis

“…Zymomonas mobilis ZM 1 (ATCC 10988) and ZM4 (ATCC 31821) were grown statically at 30 "C, in complex medium as previously described (Michel & Baratti, 1989). Growth was monitored by measurement of optical density at 450 nm.…”

“…One dimensional SDS-PAGE (10.5 YO, w/v, acrylamide) and two-dimensional PAGE were performed according to Laemmli (1970) and O'Farrell(l979, respectively. APase staining on gels was carried out as described previously (Michel & Baratti, 1989).…”

“…Zymomonas mobilis, an obligately fermentative microorganism which converts glucose, fructose and sucrose to ethanol via the Entner-Doudoroff pathway (Swings & De Ley, 1977), has been previously reported as containing a membrane-bound and phosphate-irrepressible alkaline phosphatase (Michel & Baratti, 1989). Constitutive and phosphate-irrepressible alkaline phosphatases (APases) have been described in Capnocytophaga (Poirier & Holt, 1983), Bacteroides ruminicola (Cheng & Costerton, 1973) and Bacillus licheniformis (Hulett, 1987) but these enzymes were not found associated with membranes.…”