Isolation and expression of cDNA clones encoding mammalian poly(A) polymerase

EMBO J. 1991 Dec;10(13):4251-7. doi: 10.1002/j.1460-2075.1991.tb05003.x.

Abstract

cDNA clones encoding mammalian poly(A) polymerase were isolated with probes generated by the polymerase chain reaction based on amino acid sequences derived from the purified enzyme. A bovine cDNA clone was obtained encoding a protein of 82 kDa. Expression in Escherichia coli resulted in the appearance of a poly(A) polymerase activity that was dependent on the addition of the purified specificity factor CPF and the presence of the polyadenylation signal AAUAAA in the RNA substrate. The activity copurified with a polypeptide of the expected size. A second class of cDNAs encoded a polypeptide of 43 kDa which was closely related to the N-terminal half of the 82 kDa protein. Northern blots showed two mRNAs of 4.2 and 2.4 kb that probably correspond to the two classes of cDNAs, as well as a third band of 1.3 kb. The sequence of the N-terminal half of bovine poly(A) polymerase is 47% identical with the amino acid sequence of the corresponding part of yeast poly(A) polymerase. Homologies to other proteins are of uncertain significance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Blotting, Northern
  • Cattle
  • Chromatography, Liquid
  • Cloning, Molecular
  • DNA / genetics*
  • DNA / isolation & purification
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / genetics
  • Gene Expression
  • Genes, Bacterial
  • HeLa Cells
  • Humans
  • Molecular Sequence Data
  • Plasmids
  • Polymerase Chain Reaction
  • Polynucleotide Adenylyltransferase / genetics*
  • RNA, Messenger / metabolism

Substances

  • RNA, Messenger
  • DNA
  • Polynucleotide Adenylyltransferase

Associated data

  • GENBANK/M62513
  • GENBANK/M62514
  • GENBANK/M62515
  • GENBANK/S65080
  • GENBANK/S65083
  • GENBANK/S72609
  • GENBANK/S72755
  • GENBANK/X59372
  • GENBANK/X59373
  • GENBANK/X61585