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Human DNA polymerase lambda diverged in evolution from DNA polymerase beta toward specific Mn(++) dependence: a kinetic and thermodynamic study.
- Source :
-
Biochemistry [Biochemistry] 2003 Jun 24; Vol. 42 (24), pp. 7467-76. - Publication Year :
- 2003
-
Abstract
- The recently discovered human DNA polymerase lambda (DNA pol lambda) has been implicated in translesion DNA synthesis across abasic sites. One remarkable feature of this enzyme is its preference for Mn(2+) over Mg(2+) as the activating metal ion, but the molecular basis for this preference is not known. Here, we present a kinetic and thermodynamic analysis of the DNA polymerase reaction catalyzed by full length human DNA pol lambda, showing that Mn(2+) favors specifically the catalytic step of nucleotide incorporation. Besides acting as a poor coactivator for catalysis, Mg(2+) appeared to bind also to an allosteric site, resulting in the inhibition of the synthetic activity of DNA pol lambda and in an increased sensitivity to end product (pyrophosphate) inhibition. Comparison with the closely related enzyme human DNA pol beta, as well as with other DNA synthesising enzymes (mammalian DNA pol alpha and DNA pol delta, Escherichia coli DNA pol I, and HIV-1 reverse transcriptase) indicated that these features are unique to DNA pol lambda. A deletion mutant of DNA pol lambda, which contained the highly conserved catalytic core only representing the C-terminal half of the protein, showed biochemical properties comparable to the full length enzyme but clearly different from the close homologue DNA pol beta, highlighting the existence of important differences between DNA pol lambda and DNA pol beta, despite a high degree of sequence similarity.
- Subjects :
- Allosteric Site
Base Sequence
Binding, Competitive
Catalysis
DNA Polymerase beta antagonists & inhibitors
DNA Polymerase beta chemistry
Diphosphates metabolism
Diphosphates pharmacology
Enzyme Inhibitors pharmacology
Evolution, Molecular
Humans
Kinetics
Magnesium chemistry
Magnesium pharmacology
Manganese chemistry
Manganese pharmacology
Oligonucleotides metabolism
Recombinant Proteins antagonists & inhibitors
Recombinant Proteins chemistry
Recombinant Proteins genetics
Recombinant Proteins metabolism
Thermodynamics
DNA Polymerase beta genetics
DNA Polymerase beta metabolism
Manganese metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0006-2960
- Volume :
- 42
- Issue :
- 24
- Database :
- MEDLINE
- Journal :
- Biochemistry
- Publication Type :
- Academic Journal
- Accession number :
- 12809503
- Full Text :
- https://doi.org/10.1021/bi034198m