Purification and characterization of <i>Aspergillus parasiticus</i>cytosine deaminase for possible deployment in suicide gene therapy  

作　　者：Hassan Zanna A. J. Nok S. Ibrahim H. M. Inuwa 

机构地区：[1]Department Department of Biochemistry, University of Maiduguri, Maiduguri, Nigeria [2]Department of Biochemistry, Ahmadu Bello University, Zaria, Nigeria

出　　处：《Advances in Biological Chemistry》2012年第2期152-159,共8页生物化学进展（英文）

摘　　要：Cytosine Deaminase (CD) from Aspergillus parasiticus was purified and characterized. Time course for maximal CD production (50 μmol/min/mg) was at 72 hrs. The enzyme was purified 387.73 folds with an overall yield of 13%. The CD had pH optimum of 7.2, a temperature optimum of 40℃ - 45℃, activation of energy (Ea) of 8.4 KJ/mol and a t1/2 of 1.10 hr. A. parasiticus CD stoichiometrically deaminated Cyto-sine and 5-fluorocytosine (5-FC) with the KM values of 0.19mM and 0.30 mM respectively. Studies on the effect of pH on KM and Vmax gave pKa1 of 5.8 and pKa2 of 6.3 with enthalpy of ionization of 43.01 KJ/mole suggesting histidine in the active site. The enzyme was strongly inhibited by Ca2+, Co2+, Zn2+ and Hg2+ and completely inhibited by Cu2+ and Fe2+ at 50 mM. Therefore, A. parasiticus CD can be compared with CDs from other sources that are used in suicide gene therapy.Cytosine Deaminase (CD) from Aspergillus parasiticus was purified and characterized. Time course for maximal CD production (50 μmol/min/mg) was at 72 hrs. The enzyme was purified 387.73 folds with an overall yield of 13%. The CD had pH optimum of 7.2, a temperature optimum of 40℃ - 45℃, activation of energy (Ea) of 8.4 KJ/mol and a t1/2 of 1.10 hr. A. parasiticus CD stoichiometrically deaminated Cyto-sine and 5-fluorocytosine (5-FC) with the KM values of 0.19mM and 0.30 mM respectively. Studies on the effect of pH on KM and Vmax gave pKa1 of 5.8 and pKa2 of 6.3 with enthalpy of ionization of 43.01 KJ/mole suggesting histidine in the active site. The enzyme was strongly inhibited by Ca2+, Co2+, Zn2+ and Hg2+ and completely inhibited by Cu2+ and Fe2+ at 50 mM. Therefore, A. parasiticus CD can be compared with CDs from other sources that are used in suicide gene therapy.

关 键 词：CYTOSINE DEAMINASE CYTOSINE 5-FLUOROCYTOSINE Aspergillus PARASITICUS Activation Energy 

分 类 号：R73[医药卫生—肿瘤]