title: Molecular cloning and characterization of glucose-6-phosphate dehydrogenase from Brugia malayi.
abstract: Glucose-6-phosphate dehydrogenase (G6PD), a regulatory enzyme of the pentose phosphate pathway from Brugia malayi, was cloned, expressed and biochemically characterized. The Km values for glucose-6-phosphate and nicotinamide adenine dinucleotide phosphate (NADP) were 0.25 and 0.014 mm respectively. The rBmG6PD exhibited an optimum pH of 8.5 and temperature, 40 °C. Adenosine 5' [γ-thio] triphosphate (ATP-γ-S), adenosine 5' [β,γ-imido] triphosphate (ATP-β,γ-NH), adenosine 5' [β-thio] diphosphate (ADP-β-S), Na+, K+, Li+ and Cu++ ions were found to be strong inhibitors of rBmG6PD. The rBmG6PD, a tetramer with subunit molecular weight of 75 kDa contains 0.02 mol of SH group per mol of monomer. Blocking the SH group with SH-inhibitors, led to activation of rBmG6PD activity by N-ethylmaleimide. CD analysis indicated that rBmG6PD is composed of 37% α-helices and 26% β-sheets. The unfolding equilibrium of rBmG6PD with GdmCl/urea showed the triphasic unfolding pattern along with the highly stable intermediate obtained by GdmCl.
authors.affiliation: Division of Biochemistry, CSIR-Central Drug Research Institute, Lucknow-226001, Uttar Pradesh, India.
authors.forename: Anita
authors.initials: A
authors.lastname: Verma
authors.forename: Manish K
authors.initials: MK
authors.lastname: Suthar
authors.forename: Pawan K
authors.initials: PK
authors.lastname: Doharey
authors.forename: Smita
authors.initials: S
authors.lastname: Gupta
authors.forename: Sunita
authors.initials: S
authors.lastname: Yadav
authors.forename: Prem M S
authors.initials: PM
authors.lastname: Chauhan
authors.forename: Jitendra K
authors.initials: JK
authors.lastname: Saxena
mesh_terms: D000818:Animals; D015153:Blotting, Western; D017178:Brugia malayi / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D004605:Elephantiasis, Filarial / Q000188:drug therapy; D005954:Glucosephosphate Dehydrogenase / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics; D009249:NADP / Q000378:metabolism; D018510:RNA, Helminth / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018510:RNA, Helminth; D011994:Recombinant Proteins; D009249:NADP; D005954:Glucosephosphate Dehydrogenase