Purification and characterization of glucose 6-phosphate dehydrogenase from rainbow trout (Oncorhynchus mykiss) erythrocytes

Cıftcı M., Ciltas A., Erdogan O.

Veterinarni Medicina, cilt.49, ss.327-333, 2004 (SCI-Expanded) identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Cilt numarası: 49
  • Basım Tarihi: 2004
  • Doi Numarası: 10.17221/5712-vetmed
  • Dergi Adı: Veterinarni Medicina
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus
  • Sayfa Sayıları: ss.327-333
  • Anahtar Kelimeler: purification, characterization, glucose 6-phosphate dehydrogenase, Oncorhynchus mykiss, erythrocyte, ENZYMES
  • Atatürk Üniversitesi Adresli: Evet

Özet

Glucose 6-phosphate dehydrogenase (D-glucose 6-phosphate: NADP(+) oxidoreductase, EC 1.1.1.49; G6PD) from rainbow trout (Oncorhynchus mykiss) erythrocytes was purified, using a simple and rapid method, and some characteristics of the enzyme were investigated. The purification procedure consisted of three steps: haemolysate preparation, ammonium sulphate precipitation, and 2',5'-ADP Sepharose 4B affinity gel chromatography, which took one working day. Thanks to the three consecutive procedures, the enzyme, having the specific activity of 14.51 EU/mg proteins, was purified with a yield of 70.40% and 1 271.19-fold. In order to control the purification of the enzyme SDS polyacrylamide gel electrophoresis was carried out. SDS polyacrylamide gel electrophoresis showed a single band for the enzyme. Optimal pH, stable pH, optimal temperature, molecular weight, and KM and V-max values for NADP(+) and glucose 6- phosphate (G6-P) were also determined for the enzyme. In addition, the effect of NADPH on the enzyme was investigated and K-i value and the type of inhibition were determined by means of Lineweaver-Burk graph obtained for NADPH.