Heat-labile bacterial alkaline phosphatase from a marine Vibrio sp.

Psychrophilic organisms have successfully adapted to various low-temperature environments such as cold ocean waters. Catalysts with increased catalytic efficiencies are produced, generally at the expense of thermal stability due to fewer non-covalent stabilizing interactions. A marine bacterial strain producing a particularly heat-labile alkaline phosphatase was selected from a total of 232 strains isolated from North-Atlantic coastal waters. From partial 16S rRNA sequences the strain was characterized as a Vibrio sp. An alkaline phosphatase was purified 151-fold with 54% yield from the culture medium using a single step affinity chromatography procedure on agarose-linked l-histidyldiazobenzylphosphonic acid. The active enzyme was a 55 ± 6 kDa monomer. The enzyme had optimal activity at pH 10 and was strikingly heat-labile with a half-life of 6 min at 40°C and 30 min at 32°C. This enzyme from Vibrio sp. had a higher turnover number (kcat) and higher apparent Michaelis–Menten factor (Km) than the enzyme from Escherichia coli, a clear-indication of cold-adaptation. Inorganic phosphate was a competitive inhibitor with a relatively high Ki value of 1.7 mM. Low affinity for phosphate may contribute to higher turnover rates due to more facile release of product.