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Biochemical Mechanisms of Mutagenesis and DNA Repair
Our primary research goal is to understand the molecular basis of
mutagenesis. There are 3 projects under investigation. The first is to study
biochemical and physical-chemical mechanisms governing DNA replication
fidelity. DNA polymerases make mistakes at rates as often as one in a
thousand to as little as one in a million, depending on the type of error,
surrounding DNA sequence, and on the properties of the enzyme. We have
developed a simple polyacrylamide gel electrophoresis assay to measure DNA
synthesis fidelity at any DNA template site, and we are analyzing how
fidelity depends on DNA polymerases, DNA sequences, and on protein
components of the replication complex.
When DNA is heavily damaged following exposure of a cell to chemical
mutagens or UV light, more than 20 genes in the SOS regulon of E. coli are
induced, allowing the cell to copy past nonrepairable DNA lesions resulting
in a high frequency of mutations. The second project is to investigate the
biochemical basis of SOS-induced error prone repair in E. coli. We
discovered that E. coli DNA polymerase II was induced as part of the SOS
regulon and have identified and cloned the structural gene for pol II, the
damage inducible dinA gene. Genetic and biochemical studies are currently
underway to investigate the role of pol II in vivo and to study its
properties in vitro.
Are DNA repair enzymes induced in non-dividing eucaryotic cells? The third
project is to identify and study normal and damage-induced DNA replication,
repair, and nucleotide metabolisms enzymes using neuron and astrocyte
primary and transformed cell cultures.
Selected publications
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1.
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Petruska, J., Arnheim, N., and Goodman, M. F. Stability of intrastrand
hairpin structures formed by the CAG/CTG class of DNA triplet repeats
associated with neurological diseases. Nucleic Acids Res. 24, 1992-1998
(1996).
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2.
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Bloom, L. B., Chen, X., Kuchnir Fygenson, D., Turner, J., O'Donnell, M.,
and Goodman, M. F. Fidelity of Escherichia coli DNA Polymerase III
Holoenzyme: The Effects of bets, gamma Complex Processivity Proteins and
epsilon Proofreading Exonuclease on Nucleotide Misincorporation
Efficiencies. J. Biol. Chem. 272, 27919-27930 (1997).
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3.
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Goodman, M. F. Hydrogen bonding revisited: geometric selection as a
principal determinant of DNA replication fidelity. Proc. Natl. Acad. Sci.
USA 94, 10493-10495 (1997).
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4.
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Goodman, Myron F. and Fygenson, D. F. DNA Polymerase Fidelity: From
Genetics Toward a Biochemical Understanding. Genetics 148, 1475-1482
(1998).
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5.
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Tang, M. Bruck, I., Eritja, R., Turner, J., Frank, E. G., Woodgate, R.,
O'Donnell, M., and Goodman, M. F. Biochemical Basis of SOS-Induced
Mutagenesis in Escherichia coli: Reconstitution of in vitro lesion bypass
dependent on the UmuD'2C Mutagenic Complex and RecA Protein. Proc. Natl.
Acad. Sci. USA 95, 9755-9760 (1998).
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6.
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Goodman, M. F. Purposeful Mutations. Nature 395, 221-223 (1998).
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7.
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Bertram, J. G., Bloom, L. B., Turner, J., O'Donnell, M., Beechem, J. M.,
and Goodman, M. F. Pre-steady State Analysis of the Assembly of Wild Type
and Mutant Circular Clamps of Escherichia coli DNA Polymerase III onto DNA.
J. Biol. Chem. 273, 24564-24574 (1998).
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8.
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Goodman, M. F. On the wagon: DNA polymerase joins H-bonds anonymous.
Nature Biotechnology 17, 640-641 (1999).
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9.
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Tang, M., Shen, X., Frank, E. G., O'Donnell, M., Woodgate, R., and Goodman,
M. F. UmuD'2C is an error-prone DNA polymerase, Escherichia coli pol V.
Proc. Natl. Acad. Sci. USA 96, 8919-8924 (1999).
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10.
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Rangarajan, S., Woodgate, R., and Goodman, M. F. A Phenotype For Enigmatic
DNA Polymerase II: A Pivotal Role For Pol II In Replication Restart In
UV-irradiated Escherichia coli. Proc. Natl. Acad. Sci. USA 96, 9224-9229
(1999).
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