TAILIEUCHUNG - Báo cáo khoa học: Plasmodium falciparum hypoxanthine guanine phosphoribosyltransferase Stability studies on the product-activated enzyme

Hypoxanthine guanine phosphoribosyltransferases (HGPRTs) catalyze the conversion of 6-oxopurine bases to their respective nucleotides, the phos-phoribosyl group being derived from phosphoribosyl pyrophosphate. RecombinantPlasmodium falciparumHGPRT, on purification, has negli-gible activity, and previous reports have shown that high activities can be achieved upon incubation of recombinant enzyme with the substrates hypo-xanthine and phosphoribosyl pyrophosphate [Keough DT, Ng AL, Winzor DJ, Emmerson BT & de Jersey J (1999)Mol Biochem Parasitol98, 29–41; Sujay Subbayya IN & Balaram H (2000)Biochem Biophys Res Commun 279, 433–437]. . | iFEBS Journal Plasmodium falciparum hypoxanthine guanine phosphoribosyltransferase Stability studies on the product-activated enzyme Jayalakshmi Raman Chethan S. Ashok Sujay . Subbayya Ranjith P. Anand Senthamizh T. Selvi and Hemalatha Balaram Molecular Biology and Genetics Unit JawarharlalNehru Centre for Advanced Scientific Research Jakkur Bangalore India Keywords active state stability hypoxanthine guanine phosphoribosyltransferase Plasmodium falciparum product activation thermal stability Correspondence H. Balaram Molecular Biology and Genetics Unit JawarharlalNehru Centre for Advanced Scientific Research Jakkur Bangalore 560064 India Fax 91 80 22082766 Tel 91 80 22082812 E-mail hb@ Received 20 October 2004 revised 14 February 2005 accepted 18 February 2005 doi Hypoxanthine guanine phosphoribosyltransferases HGPRTs catalyze the conversion of 6-oxopurine bases to their respective nucleotides the phosphoribosyl group being derived from phosphoribosyl pyrophosphate. Recombinant Plasmodium falciparum HGPRT on purification has negligible activity and previous reports have shown that high activities can be achieved upon incubation of recombinant enzyme with the substrates hypoxanthine and phosphoribosyl pyrophosphate Keough DT Ng AL Winzor DJ Emmerson BT de Jersey J 1999 Mol Biochem Parasitol 98 29-41 Sujay Subbayya IN Balaram H 2000 Biochem Biophys Res Commun 279 433-437 . In this report we show that activation is effected by the product Inosine monophosphate IMP and not by the substrates. Studies carried out on Plasmodium falciparum HGPRT and on a temperaturesensitive mutant L44F show that the enzymes are destabilized in the presence of the substrates and the product IMP. These stability studies suggest that the active product-bound form of the enzyme is less stable than the ligand-free unactivated enzyme. Equilibrium isothermal-unfolding studies indicate that the active form is destabilized by 2-3 kcal-mol-1 compared

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