Title

Increasing the yield of Glucose-6-Phosphate Dehydrogenase.

Document Type

Article

Publication Date

2001

Abstract

Glucose-6-Phosphate Dehydrogenase (G6PD) is responsible for catalyzing the initial oxidative reaction- the conversion of Glucose-6-Phosphate (G6P) into 6-Phosphogluconolactone - in the Pentose Phosphate Pathway (PPP). It has been discovered that for every mole of glucose-6-phosphate entering PPP, one mole of NADPH and a five-carbon sugar (ribose) is produced. NADPH, utilized in cellular biosynthetic reductive reactions, is found to be the most significant pathway in the mature erythrocytes (red blood cells) for the production of reduced glutathione that regulates the oxidative stress within red blood cells. The deficiency in the activity of the Glucose-6-phosphate dehydrogenase, an X-chromosomal linked enzyme, is suggested to be associated with numerous variants of the enzyme that trigger the resistance to malarial parasite, Plasmodium falciparum . Various mutations of the enzyme are found in high frequency in Asiatic, African, and Mediterranean populations. Since the 1950's, studies have been conducted to examine variants and elucidate biochemical, biological, and genetic properties of the enzyme. Nevertheless, the complications associated with purifying appropriate amounts of enzyme have always been a major problem for research projects examining the nature of glucose-6-phosphate dehydrogenase. Current study has been aimed at elevating the yields of the human recombinant enzyme by employing methods for increasing the growth of Escherichia coli cells from which the enzyme is extracted and investigating the direct relationship between cell concentration and activity of the G6PD. Moreover, freeze/thaw techniques have been applied for reducing the degree of difficulty and improving the effectiveness of rupturing cells (sonication), and affi-gel blue gel chromatography was used to further isolate and stabilize the enzyme.

Advisor

Chris L. Craney

Department

biochem

Support

NSF-Research Experience for Undergraduates grant



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