Mammalian DNA ligases. Catalytic domain and size of DNA ligase I.
DNA ligase I is the major DNA ligase activity in proliferating mammalian cells. The protein has been purified to apparent homogeneity from calf thymus. It has a monomeric structure and a blocked N-terminal residue. DNA ligase I is a 125-kDa polypeptide as estimated by sodium dodecyl sulfate-gel electrophoresis and by gel chromatography under denaturing conditions, whereas hydrodynamic measurements indicate that the enzyme is an asymmetric 98-kDa protein. Immunoblotting with rabbit polyclonal antibodies to the enzyme revealed a single polypeptide of 125 kDa in freshly prepared crude cell extracts of calf thymus. Limited digestion of the purified DNA ligase I with several reagent proteolytic enzymes generated a relatively protease-resistant 85-kDa fragment. This domain retained full catalytic activity. Similar results were obtained with partially purified human DNA ligase I. The active large fragment represents the C-terminal part of the intact protein, and contains an epitope conserved between mammalian DNA ligase I and yeast and vaccinia virus DNA ligases. The function of the N-terminal region of DNA ligase I is unknown.
Imperial Cancer Research Fund, Clare Hall Laboratories, South Mimms, Hertfordshire, United Kingdom.
A E
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Tomkinson
D D
DD
Lasko
G
G
Daly
T
T
Lindahl
D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D002417:Cattle; D003429:Cross Reactions; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D000939:Epitopes; D006801:Humans; D008970:Molecular Weight; D010449:Peptide Mapping; D011118:Polynucleotide Ligases / Q000378:metabolism*; D013381:Subtilisins / Q000494:pharmacology; D013950:Thymus Gland / Q000201:enzymology; D014461:Ultracentrifugation
D000939:Epitopes; C000607892:LIG1 protein, human; D013381:Subtilisins; D011088:DNA Ligases; D011118:Polynucleotide Ligases; D000072481:DNA Ligase ATP