الفهرس 
Preparation of sephacryl S-300 columnOnly 14 pages are availabe for public view
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Abstract
Five glucose-6-phosphate dehydrogenase (G6PD) isoenzymes
were purified to homogeneity from the liver of the buffalo, camel and
sheep. The purification procedures involved crude extraction,
ammonium sulfate precipitation and anion exchange
chromatography on DEAE-cellulose column, gel filtration through
sephacryl S-300 column and chromatography on 2′, 5′ ADP
Sepharose 4B affinity column. All of the purified isoenzymes turned
out to be homogeneous as judged by native PAGE.
The buffalo G6PD (BLG6PD1 and BLG6PD2) have molecular
weights of 28 kDa and 65 kDa. They have pI values of 7.7-7.9 and
5.7- 5.9 and displayed their maximum activity at pH 8.0 and 8.2
respectively. The Km values were found to be 0.059 and 0.06 mM
NADP+ for BLG6PD1 and BLG6PD2. NADPH is found to be a
competitive inhibitor for both isoenzymes.
The camel G6PD (CLG6PD) has molecular weight of 194 kDa,
pI value of 6.6-6.8 and displayed its maximum activity at pH 7.8.
CLG6PD has Km value of 0.081 mM NADP+ and inhibited
competitively with NADPH.
The sheep G6PD (SLG6PD1 and SLG6PD2) have molecular
weights of 177 kDa and 65 kDa. They have pI values of 5.9-6.1 and
5.4-5.6 and displayed their maximum activity at pH 7.8 and 8.0
respectively. The Km values were found to be 0.079 and 0.058 mM
NADP+ for SLG6PD1 and SLG6PD2. NADPH is found to be a
competitive inhibitor for both isoenzymes.
The purified buffalo BLG6PD1 is used in preparation of
glucose hexokinase diagnostic kit. The prepeared kit is linear, stable
and sensitive in determinination of glucose concentrations and
compared with commercially available kit (sigma glucose hexokinase
diagnostic kit) using diabetic patient samples.
Key words: Glucose-6-phosphate dehydrogenase, purification and
characterization, buffalo, camel and sheep liver.