Biochemical Studies of Starch Phosphorylase from Rice Grains

• Main Authors: HSU, Ren-Hung, 許仁弘
• Other Authors: Jong-Ching SU, Ping-Du LEE
• Format: Others
• Language: zh-TW
• Published: 1997
• Online Access: http://ndltd.ncl.edu.tw/handle/55206401755184917352

碩士 === 國立臺灣大學 === 農業化學系 === 85 === Degraded fragments of H form starch phosphorylase (SP) of the milky rice (Oryza sativa) grains are purified by ammonium sulfate fractionation, DEAE-Sepharose CL-6B anion exchange chromatography, Con A-Sepharose affinity chromatography and Superose 12 fast protein liquid chromatography. Up to the step of affinity chromatography, the purification fold is 34; the enzyme activity recovery is about 5%. No SP isozyme was found at the unbound fractions after DEAE- Sepharose CL-6B chromatography. By using activity staining on 7.5% native-PAGE, it was found that there is difference of electrophoretic mobility between the fractions obtained from anion-exchange and affinity chromatography. It was bounded by the Con A-Sepharose affinity chromatography without polysaccharide as ligand, so SP is a glycoprotein. The subunit molecular mass of SP is about 90 kD. Being determined by SDS-PAGE, the molecular mass of the degraded SP after Superose 12 (FPLC) was about 66 kD. The molecular mass of degraded SP eluted from affinity preparative electrophoresis obtained on 15% and 12.5% SDS-PAGE are 66 kD and 30 kD. The optimum and most stable pH for SP all are 5.4, and the optimum reaction temperature is about 40℃. The rice grain SP has higher affinity with glycogen, soluble starch and dextrin rather than that with maltooligosaccharide.