RESUMEN
A species of DNA polymerase alpha that is active in the ATP(GTP)-dependent conversion of MVM parvovirus single-stranded linear DNA to the duplex replicative form has been purified 4300-fold from Ehrlich ascites mouse tumour cells. The single-stranded----replicative form activity is maintained throughout ammonium sulfate precipitation, DEAE-cellulose, phosphocellulose and hydroxyapatite column chromatography and glycerol gradient sedimentation. Polypeptides with Mr = 230 000, 220 000, 183 000, 157 000, 125 000, 70 000, 65 000, 62 000, 57 000, 53 000 and 48 000 copurify with the single-stranded----replicative form activity, which sediments at approx. 10 S. The Mr = 183 000, 157 000 and 125 000 polypeptides exhibit catalytic activity when assayed in situ following SDS-polyacrylamide gel electrophoresis. The 10 S form of DNA polymerase alpha is functionally distinguishable from an 8.4 S form of the enzyme obtained from the same cells on the basis of single-stranded----replicative form activity. The single-stranded----replicative form activity of the 10 S enzyme is stable at 22 degrees C for up to 3 h, but exhibits a half life of only 5 min at 45 degrees C.
Asunto(s)
Adenosina Trifosfato/farmacología , Carcinoma de Ehrlich/enzimología , ADN Polimerasa II/metabolismo , Replicación del ADN/efectos de los fármacos , ADN de Cadena Simple/genética , ADN Viral/genética , Guanosina Trifosfato/farmacología , Virus Diminuto del Ratón/genética , Parvoviridae/genética , Animales , ADN Polimerasa II/aislamiento & purificación , Electroforesis en Gel de Poliacrilamida , Ratones , Peso MolecularRESUMEN
At different exponential growth rates obtained either by varying the carbon source of the culture medium or limiting glucose uptake, intracellular levels of putrescine and spermidine were measured. Over a ten-fold increase in growth rate an approximately three-fold increase in putrescine level and a 3.5-fold increase in spermidine level per cell absorbance were observed. Conditions favoring an abrupt alteration in growth rate, such as occur following nutritional shiftup of Escherichia coli, resulted in a significant increase in the intracellular level of putrescine and virtually no change in the spermidine level. Because of the magnitude and the timing of the change in polyamine levels, the hypothesis that polyamines are (the components) responsible for inducing the rapid increase in the rate of RNA synthesis following nutritional shiftup is rejected.
