| Popis: |
We studied deoxyadenosine and deoxyguanosine phosphorylating activities in human placental cytosol. The specific activities of nucleoside kinase enzymes in nanomoles per h per mg +/- SD were as follows: adenosine kinase, 30 +/- 14; deoxyadenosine kinase, 12 +/- 2; deoxycytidine kinase, 0.30 +/- 0.04; and deoxyguanosine kinase, 27 +/- 16. Three major activities were resolved by ion exchange and affinity chromatography: deoxyguanosine-deoxycytidine kinase, deoxycytidine-deoxyadenosine kinase, and adenosine-deoxyadenosine kinase. Two other activities contained significant quantities of deoxyadenosine kinase. Deoxyguanosine-phosphorylating activity eluted as a single peak in association with deoxycytidine kinase. This deoxyguanosine-deoxycytidine kinase had an apparent molecular weight of 54,000, a Stokes radius of 31 A, and apparent Km values of 10, 130, and 14 microM for deoxyguanosine, deoxycytidine, and ATP, respectively. Four peaks of deoxyadenosine phosphorylating activity were resolved by affinity chromatography with AMP-Sepharose 4B. Adenosine-deoxyadenosine kinase had an apparent molecular weight of 38,000, a Stokes radius of 27.4 A, and apparent Km values of 0.4, 510, and 75 microM for adenosine, deoxyadenosine, and ATP, respectively. Attempts to distinguish whether adenosine-deoxyadenosine kinase was one enzyme with these two activities or two separate enzymes suggested that the former was the case. Deoxycytidine-deoxyadenosine kinase had apparent Km values of 0.7, 670, and 12 microM for deoxycytidine, deoxyadenosine, and ATP, respectively. Its apparent molecular weight was estimated to be 49,000 and its Stokes radius 30 A. Two other minor peaks of deoxyadenosine-phosphorylating activity had characteristics different from either deoxycytidine kinase or adenosine kinase-associated deoxyadenosine kinase. Our studies indicate that human placental cytosol contains a complex mixture of nucleoside kinase enzymes. |
