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 degrees C and 30 min at 32 degrees C. This enzyme from Vibrio sp. had a higher turnover number (k(cat)) and higher apparent Michaelis-Menten factor (K(m)) than the enzyme from Escherichia coli, a clear-indication of cold-adaptation. Inorganic phosphate was a competitive inhibitor with a relatively high K(i) value of 1.7 mM. Low affinity for phosphate may contribute to higher turnover rates due to more facile release of product.
Hauksson
JB
Department of Chemistry, Science Institute, University of Iceland, Dunhaga 3, 107, Reykjavik, Iceland
Andrésson
OS
Ásgeirsson
B