Structure of rat DNA polymerase beta revealed by partial amino acid sequencing and cDNA cloning. | Zendy

Barbara Z. Zmudzka | Zendy; Debomita Sengupta | Zendy; Akio Matsukage | Zendy; Fabio Cobianchi | Zendy; Pradeep G. Kumar | Zendy; Samuel H. Wilson | Zendy

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Structure of rat DNA polymerase beta revealed by partial amino acid sequencing and cDNA cloning.

Author(s) -

Barbara Z. Zmudzka,

Debomita Sengupta,

Akio Matsukage,

Fabio Cobianchi,

Pradeep G. Kumar,

Samuel H. Wilson

Publication year - 1986

Publication title -

proceedings of the national academy of sciences

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 5.011

H-Index - 771

eISSN - 1091-6490

pISSN - 0027-8424

DOI - 10.1073/pnas.83.14.5106

Subject(s) - complementary dna , microbiology and biotechnology , biology , cdna library , peptide sequence , open reading frame , amino acid , nucleic acid sequence , messenger rna , polymerase , rapid amplification of cdna ends , rna , dna polymerase , protein primary structure , dna , biochemistry , gene

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.

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