Limited proteolysis of DNA polymerases as probe of functional domains.
Various proteolytic enzymes have been used to probe for domains in DNA polymerases. Results with several DNA polymerases that have been subjected to partial proteolysis demonstrated that there is a modular organization with different activities located in separate domains. In the case of the Klenow fragment, these domains appear to be independent of each other. With other DNA polymerases, the question of modular independence is not settled. Limited proteolysis for probing structure has been used with many other proteins in addition to DNA polymerases and the information obtained has been helpful in interpreting function-structure relationships. It is a general approach and can be applied in situations where the existence of domains is suspected. The simplicity of the method and the ease of monitoring the outcome is probably the main reason for its widespread and increasing use in enzymology.
Konigsberg
W H
WH
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06510, USA.
D002918:Chymotrypsin / Q000378:metabolism; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004259:DNA-Directed DNA Polymerase / Q000737:chemistry* / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D010450:Endopeptidases / Q000378:metabolism*; D004926:Escherichia coli; D006868:Hydrolysis; D007700:Kinetics; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D013816:Thermodynamics; D014764:Viral Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D018076:DNA, Complementary; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014764:Viral Proteins; C069983:gene 43 protein, Enterobacteria phage T4; D004259:DNA-Directed DNA Polymerase; D010450:Endopeptidases; D002918:Chymotrypsin