Structure of rat DNA polymerase beta revealed by partial amino acid sequencing and cDNA cloning.
A cDNA library of newborn rat brain poly(A)+ RNA in phage lambda gt11 was screened with a polyclonal antibody against chicken DNA polymerase beta. One positive phage was isolated and purified after testing 2 X 10(7) recombinants. This phage, designated lambda pol beta-10, contained an 1197-base-pair cDNA insert that corresponded to a mRNA with a poly(A) sequence at the 3' terminus and a single, long open-reading frame of 957 bases. The open-reading frame, starting 44 residues from the 5' end of the cDNA, predicted a 36,375-Da protein of 318 amino acids. Comparison of this deduced amino acid sequence with the partial sequence obtained with purified polymerase beta revealed a match of six tryptic peptides, involving a total of 47 amino acid residues. This confirmed the identity of the cDNA. Blot-hybridization analysis of newborn rat brain poly(A)+ RNA revealed a mRNA species of approximately the same size as the cDNA insert; in addition, a second mRNA species approximately equal to 4000 bases long was detected. Computer-derived secondary structure analysis of the enzyme predicted seven regions of alpha-helix distributed throughout and three regions of beta-sheet.
Zmudzka
B Z
BZ
SenGupta
D
D
Matsukage
A
A
Cobianchi
F
F
Kumar
P
P
Wilson
S H
SH
D000595:Amino Acid Sequence; D000818:Animals; D002645:Chickens / Q000235:genetics; D003001:Cloning, Molecular; D004247:DNA / Q000032:analysis; D004256:DNA Polymerase I / Q000235:genetics*; D011061:Poly A / Q000235:genetics; D011487:Protein Conformation; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D012689:Sequence Homology, Nucleic Acid
D012333:RNA, Messenger; D011061:Poly A; D004247:DNA; D004256:DNA Polymerase I