Purification and Characterization of Mannitol-l-Phosphatase in the Red Alga Caloglossa continua (Ceramiales, Rhodophyta)

Purification of mannitol-l-phosphatase, an enzyme catalyzing the final step of mannitol biosynthesis, was first achieved in the mannitol-accumulating red alga Caloglossa continua (Okamura) King et Puttock. The enzyme was shown to be a monomer, since gel filtration and sodium dodecyl sulfate–polyacrylamide gel electrophoresis gave close values of apparent molecular weights of 28,500 and 30,200, respectively. The protein exhibited an isoelectric point of 4.8. The substrate specificity for mannitol-l-phosphate (MIP) was very high, and that for K _m(MIP) was 0.41 mM. The catalytic activity was optimal at pH 7.4. The enzyme was activated by Mg²⁺, but was strongly inhibited by Ca²⁺, NaF, N-ethylmaleimide, and p-hydroxymercuribenzoic acid. Seawater levels of NaCl and physiological levels of mannitol also inhibited the activity by 50% or more. Changes in the concentrations of those ions and metabolites may regulate the biosynthesis of mannitol as an osmoregulant in vivo. Received May 7, 2001; accepted June 15, 2001.