Adeyinka Afolayan

Purification and characterization of glutathione transferase from the giant African snail, Archachatina marginata

1. Glutathione-S-transferase has been purified from the hepatopancreas of Archachatina marginata to homogeneity. 2. The enzyme was found to be a dimer with a molecular weight of 44,000. The subunits sizes were 22,500 and 23,500 respectively. The isoelectric points of the enzyme were 8.35, 7.95 and 4. The enzyme was most stable at temperature below 40 degrees C. Upon denaturation by 4 M urea, only 56% of the activity could be recovered. 3. The Kms for glutathione and 1-chloro-2,4-dinitrobenze (CDNB) were 0.23 mM and 0.4 mM respectively. The specific activity of the enzyme with CDNB and p-nitrophylacetate as substrates were 47 mumol/mg and 38 mumol/mg respectively. 4. Inhibition studies showed that S-hexylglutathione, Rose Bengal, iodoacetamide, sodium azide and Procion Blue H-B were good inhibitors with I50 values ranging from 18.5 microM to 299 mM. 5. The amino acid composition showed that the enzyme had a relatively high content of hydrophobic and acidic amino acid residues. The peptide maps of the tryptic digests of the native and performic acid-oxidised enzyme indicated that there might be about two disulphide bridges per molecule of the enzyme.

Isolation and properties of creatine kinase from the breast muscle of tropical fruit bat, Eidolon helvum (kerr)

Creatine kinase, from fruit bat breast muscle, has been purified to homogeneity. The mol. wt of the enzyme was estimated to be about 78,000-80,000 with two subunits of 42,500. There are nine thiol residues/mol of the enzyme and two of these react readily with DTNB leading to total inactivation of the enzyme. The metal ion specificity was in order Mg2+ greater than Zn2+ greater than Co2+. Initial velocity and product inhibition studies of the reverse reaction are consistent with sequential reaction but of either rapid equilibrium random or ordered type.

Purification and some properties of pyruvate kinase from the skeletal muscle of African land tortoise Kinixys erosa (Linn)

Abstract 1. 1. Pyruvate kinase from Africa land tortoise (Kinixys erosa) skeletal muscle was isolated and purified to homogeneity. 2. 2. The mol. wt of the enzyme was estimated to be 212,333 ± 2887 with four subunits of 49,680 ± 526 . 3. 3. The enzyme, denatured by 4M guanidine-HCl, regained a maximum of 80–87% of its original activity upon dilution at 20°C and at a protein concentration of 80 μg/ml in appropriate buffer containing 10 mM PEP and 1 mM l -valine. The kinetics of renaturation was first order. 4. 4. The catalytically active renatured enzyme was a dimer even though it was kinetically similar to the tetrameric native enzyme.

The plasma membrane of human erythrocyte with different levels of glucose-6-phosphate dehydrogenase

Abstract 1. 1. The erythrocyte ghosts from red cells having normal or deficient level of glucose-6-phosphate dehydrogenase have been prepared. 2. 2. The two types of ghosts appear similar in the size and distribution of polypeplides and glyco-proteins. 3. 3. However, chemical modification and fluorescence studies in the presence of primaquine and chloroquine suggest a possible difference in structure between the two types of ghost. 4. 4. Generally, the ghost from red cell with defficient G6PD is more perturbed by antimalarial drugs. 5. 5. This result may explain why carriers of this deficient enzyme have their red cells readily haemolysed on consumption of antimalarial drugs.