Identification and characterization of human DNA polymerase beta 2, a DNA polymerase beta -related enzyme.
The BRCA1 COOH terminus (BRCT) motif is present in many nuclear proteins that contribute to cell cycle regulation or DNA repair. Polymerase chain reaction-based screening with degenerate primers targeted to the BRCT motif resulted in the isolation of a human cDNA for a previously unidentified DNA polymerase (designated DNA polymerase beta2) that is closely related to DNA polymerase beta (Pol beta). The predicted Pol beta2 protein contains a BRCT motif in its NH(2)-terminal region; its COOH-terminal region exhibits 33% sequence identity to a corresponding region of human Pol beta. The Pol beta2 gene is expressed in a tissue-specific manner, with transcripts being most abundant in testis. A fusion construct comprising Pol beta2 and green fluorescent protein exhibited a predominantly nuclear localization in transfected HeLa cells. Recombinant human Pol beta2 from insect cells exhibited substantial DNA polymerase activity, but it did not possess terminal deoxyribonucleotidyl transferase activity. A truncated Pol beta2 mutant lacking the BRCT motif retained substantial DNA polymerase activity, whereas a mutant Pol beta2 with two alanine point mutations within the DNA polymerase active site did not. These results indicate that Pol beta2 is a Pol beta-related DNA polymerase with a BRCT motif that is dispensable for its polymerase activity.
Nagasawa
K
K
Department of Geriatric Research, National Institute for Longevity Sciences, Obu, Aichi 474-8522, Japan.
Kitamura
K
K
Yasui
A
A
Nimura
Y
Y
Ikeda
K
K
Hirai
M
M
Matsukage
A
A
Nakanishi
M
M
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D015152:Blotting, Northern; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D004253:DNA Nucleotidylexotransferase / Q000378:metabolism; D019951:DNA Polymerase beta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D017404:In Situ Hybridization, Fluorescence; D007313:Insecta; D008164:Luminescent Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014018:Tissue Distribution
D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; C415367:DNA polymerase beta2; D004253:DNA Nucleotidylexotransferase; D019951:DNA Polymerase beta