This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Moss, B. Articles by Rosenblum, E. N. Search for Related Content PubMed PubMed Citation Articles by Moss, B. Articles by Rosenblum, E. N.

Vaccinia Virus Polyriboadenylate Polymerase: Covalent Linkage of the Product with Polyribonucleotide and Polydeoxyribonucleotide Primers

¹ Laboratory of Biology of Viruses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20014

ABSTRACT

A polyriboadenylate [poly(A)] polymerase, purified from vaccinia virus cores, was stimulated by polydeoxyriboadenylate: polydeoxyribothymidylate [poly(dA:dT)] and by polyribocytidylate [poly(C)] primers suggesting mechanisms of either transcription or terminal addition. Evidence for the latter was obtained by the demonstration of covalent linkages between the poly(A) products and both primers. In 99% dimethylsulfoxide-sucrose gradients, the sedimentation of poly(A) formed with poly(dA: dT) primer was reduced after DNase I treatment and the sedimentation of poly(A) formed with poly(C) primer was reduced by RNase A treatment, whereas the sedimentation of poly(A) formed without primer was not affected by either. Formation of a phosphodiester bond between primer and product was demonstrated by means of isotope transfer experiments. ³²P from -[³²P]ATP was transferred to 2'(3')-CMP after alkaline or enzymatic hydrolysis of the poly(C)-primed polymerase reaction product. Transfer primarily or exclusively to 3'-dTMP was found after enzymatic hydrolysis of the poly(dA: dT)-primed polymerase reaction product. The elution pattern of the poly(A) polymerase from DNA-cellulose suggested that a single enzyme catalyzes the attachment of adenylate residues to both polyribonucleotide and polydeoxyribonucleotide primers; nevertheless the purest enzyme preparations contain two bands resolved by polyacrylamide gel electrophoresis in sodium dodecyl sulfate.