Biochemical characterization of mutant forms of DNA polymerase I from Escherichia coli. I. The polA12 mutation.
DNA polymerase I has been purified to greater than 90% homogeneity from a strain of Escherichia coli K12 that bears the temperature-sensitive DNA polymerase I mutatation, polA12. The mutant enzyme has a reduced electrophoretic mobility and sedimentation rate. It is abnormally thermolabile and is rapidly inactivated at low salt concentrations. Its polymerase and 5' leads to 3' exonuclease activities are not grossly defective at 30 degrees, yet its capacity to promote the concerted 5' leads to 3' polymerization and the 5' leads to 3' exonucleolytic hydrolysis of nucleotides at a nick ("nick translation") is decreased 10-fold. These effects are probably the result of a significant alteration in the tertiary structure of the enzyme.
Biochemical characterization of mutant forms of DNA polymerase I from Escherichia coli. I. The polA12 mutation. DNA polymerase I has been purified to greater than 90% homogeneity from a strain of Escherichia coli K12 that bears the temperature-sensitive DNA polymerase I mutatation, polA12. The mutant enzyme has a reduced electrophoretic mobility and sedimentation rate. It is abnormally thermolabile and is rapidly inactivated at low salt concentrations. Its polymerase and 5' leads to 3' exonuclease activities are not grossly defective at 30 degrees, yet its capacity to promote the concerted 5' leads to 3' polymerization and the 5' leads to 3' exonucleolytic hydrolysis of nucleotides at a nick ("nick translation") is decreased 10-fold. These effects are probably the result of a significant alteration in the tertiary structure of the enzyme.