Assignments of 1H, 15N, and 13C resonances for the backbone and side chains of the N-terminal domain of DNA polymerase beta. Determination of the secondary structure and tertiary contacts.
DNA polymerase beta consists of an N-terminal single-stranded DNA binding domain and a C-terminal catalytic domain separable by mild proteolysis [Kumar et al. (1990) J. Biol. Chem. 265, 2124-2131]. The N-terminal domain participates in template and gapped DNA recognition and contributes significantly to catalysis. The secondary structure and tertiary contacts within the cloned N-terminal domain (residues 2-87) of mammalian DNA polymerase beta have been determined using multidimensional NMR. Assignments of backbone 1H, 15N, and 13C resonances and side chain 1H and 13C resonances have been obtained from double- and triple-resonance 3D NMR experiments. The 13C-edited TOCSY experiment has allowed nearly complete assignments of 1H and 13C resonances within side chains. The 13C-edited NOESY experiment has been used for determination of medium- and long-range NOEs and a determination of tertiary contacts. The N-terminal domain is found to consist of four helices, helix-1 (15-26), helix-2 (36-47), helix-3 (56-61), and helix-4 (69-78), which on the basis of long-range NOEs are tightly packed of form a hydrophobic core. The remainder of the domain consists of two turns (48-51 and 62-65), an omega-type loop (27-35), and extended structure. The aromatic side chains of Y36, Y39, Y49, and F76 display tertiary contacts indicative of at least partial hydrophobic packing. The S30 and H34 residues which cross-link to single-stranded DNA [Prasad et al. (1993) J. Biol. Chem. 268, 15906-15911] are contained within the K27-K35 loop.(ABSTRACT TRUNCATED AT 250 WORDS)
Liu
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Department of Chemistry, University of Wisconsin-Milwaukee 53211.
DeRose
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Prasad
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Wilson
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Mullen
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D000595:Amino Acid Sequence; D000818:Animals; D004253:DNA Nucleotidylexotransferase / Q000737:chemistry; D004256:DNA Polymerase I / Q000737:chemistry* / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid
D010446:Peptide Fragments; D011994:Recombinant Proteins; C089040:DNA polymerase I (2-87); D004253:DNA Nucleotidylexotransferase; D004256:DNA Polymerase I