Purification and characterization of trehalose-6-phosphate synthase from Saccharomycopsis fibuligera A11.
Mutant A11, a mutant of Saccharomycopsis fibuligera Sdu with low acid and neutral trehalase was found to accumulate over 18% (w/w) trehalose from starch in its cells. In this study, trehalose-6-phosphate synthase (Tps1) was purified to homogeneity from this mutant, with a 30-fold increase in the specific enzyme activity, as compared to the concentrated cell-free extract, from initial cells. The molecular mass of the purified enzyme as determined by SDS-PAGE was 66 kDa. The optimum pH and temperature of the purified enzyme were 6.6 and 37 degrees C, respectively. The enzyme was activated by Ca2+, K+ and Mg2+, with K+ showing the highest activation at 35 mM. On the other hand, Mn2+, Cu2+, Fe3+, Hg2+ and Co2+ inhibited the enzyme. The enzyme was also strongly inhibited by protease inhibitors such as iodoacetic acid, EDTA and PMSF.
Liang
Likun
L
UNESCO Chinese Center of Marine Biotechnology, Ocean University of China, Yushan Road, No.5, Qingdao, China.
Chi
Zhenming
Z
Gao
Lingmei
L
Ma
Liyan
L
D002474:Cell-Free System; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004492:Edetic Acid / Q000737:chemistry; D005964:Glucosyltransferases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D019807:Iodoacetic Acid / Q000737:chemistry; D007477:Ions; D009154:Mutation; D011480:Protease Inhibitors / Q000494:pharmacology; D012442:Saccharomycopsis / Q000201:enzymology*; D013696:Temperature; D014199:Trehalose / Q000737:chemistry
D007477:Ions; D011480:Protease Inhibitors; D004492:Edetic Acid; D014199:Trehalose; D005964:Glucosyltransferases; C023180:trehalose-6-phosphate synthase; D019807:Iodoacetic Acid