EXTRACTION, ASSAY AND SOME PROPERTIES OF FLORIDOSIDE PHOSPHATE SYNTHASE FROM PORPHYRA-PERFORATA (RHODOPHYTA)

A suitable method for extraction of floridoside phosphate synthase (FPS, UDP-galactose: sn-3-glycerol phosphate: 1 --> 2' alpha-D-galactosyl transferase) from Porphyra perforata J. Ag. was developed. Two assay methods for enzyme activity were utilized, one measuring the amount of floridoside formed by using gas-liquid chromatography, the other measuring the sn-3-glycerol phosphate-dependent formation of UDP; both assays gave similar results. FPS is a soluble protein, and FPS activity in the extract as determined by the amount of product formed in vitro compared well with the in vivo rate of floridoside synthesis (4-7-mu-mol product formed.h-1.g-1 fresh wt). The rate of product formation in vitro was linear up to 45 min and proportional to protein concentration in the assay mixture. The temperature optimum was 30-35-degrees-C. FPS was active over a range of pH values from 7.0-8.5. It was stable in concentrated solutions in the presence of 0.3 M ammonium sulfate, but activity was lost in diluted solution (protein concentration below 0.2 mg.mL-1) or below 0.2 M ion strength. The data suggest that FPS may be an oligomeric protein which occurs free in the cytoplasm or loosely bound to a membrane. It may also be a regulatory protein controlling the overall rate of synthesis of floridoside in vivo.