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Scientific Overview Research Interest Summary Principal Investigators    Yuri Bushkin, Ph.D.
   Loren Day, Ph.D.
   Karl Drlica, Ph.D.
   David Dubnau, Ph.D.
   Marila Gennaro, M.D.
   Gilla Kaplan, Ph.D.
   Fred Kramer, Ph.D.
   Barry Kreiswirth, Ph.D.
   Leonard Mindich, Ph.D.
   Harvey Penefsky, Ph.D.
   David Perlin, Ph.D.
   Richard Pine, Ph.D.
   Abraham Pinter, Ph.D.
   Issar Smith, Ph.D.
   Patricia Soteropoulos, Ph.D.
   Sanjay Tyagi, Ph.D.
   David Wah, Ph.D.
   Shuishu Wang, Ph.D.

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Junior Faculty Members Research Grants
 
Alexander Goldfarb, Ph.D.

Research Summary  |  Recent Articles  |  C.V.
 

Research Summary

RNA polymerase is the central enzyme of gene expression and the target for genetic regulation. We are interested in RNA polymerase from bacterium Escherichia coli and its regulation in structure-functional terms. To this end, we have developed a unique experimental system which is based on two technical advances: (i) the assembly of functional RNA polymerase from individually over expressed subunits, and (ii) the development of a genetic system for in vitro mutagenesis of rpo genes and in vivo maintenance of dominant lethal mutants. This system makes it possible, for the first time, to attack vital functions of TNA polymerase using the combination of genetic, physiological and biochemical analyses. The choice of sites for site-directed mutagenesis of rpo genes is dictated by (i) homology alignment of RNA polymerase subunits from different organisms which reveals localities that are highly conserved in evolution; and (ii) mapping of functionally important sites by chemical derivatization with specific affinity reagents, such as crosslinkable analogs of nucleotide substrates and RNA transcripts.

The second line of research is concerned with in vitro mechanism of action of regulatory factors that act on RNA polymerase and modulate its function. We have demonstrated that Lac repressor acts by preventing promoter clearance from a joint complex with RNA polymerase. In another project, we have discovered a novel family of transcriptional factors in E. coli, that act at the elongation stage by helping RNA polymerase to resume transcription from the so-called "elongation-arresting sites". Such sites occasionally trap elongating transcriptional complexes resulting in permanent stalling of RNA polymerase. The two factors that we found, TcfA and TcfB, act independently by cleaving off the tip of the nascent transcript in the arrest complex, allowing RNA polymerase to back up a retry reading through the potential elongation trap.




Recent Articles

Mustaev A, Zaychikov E, Grachev M, Kozlov M, Severinov K, Epshtein V, Korzheva N, Bereshchenko O, Markovtsov V, Lukhtanov E, Tsarev I, Maximova T, Kashlev M, Bass I, Nikiforov V, Goldfarb A. (2003).
Strategies and methods of cross-linking of RNA polymerase active center.
Methods Enzymol. 2003;371:191-206
PMID: 14712701

Sosunova E, Sosunov V, Kozlov M, Nikiforov V, Goldfarb A, Mustaev A. (2003).
Donation of catalytic residues to RNA polymerase active center by transcription factor Gre.
Proc Natl Acad Sci U S A. 2003 Dec 23;100(26):15469-74. Epub 2003 Dec 10
PMID: 14668436
Abstract  |  Full Article

Sosunov V, Sosunova E, Mustaev A, Bass I, Nikiforov V, Goldfarb, A. (2003).
Unified two-metal mechanism of RNA synthesis and degradation by RNA polymerase.
EMBO J. 2003 May 1;22(9):2234-44
PMID: 12727889
Abstract  |  Full Article

Epshtein V, Mustaev A, Markovtsov V, Bereshchenko O, Nikiforov V, Goldfarb A. (2002).
Swing-gate model of nucleotide entry into the RNA polymerase active center.
Mol Cell 2002 Sep;10(3):623-34
PMID: 12408829
Abstract  |  Full Article

Campbell EA, Korzheva N, Mustaev A, Murakami K, Nair S, Goldfarb A, Darst SA. (2001).
Structural mechanism for rifampicin inhibition of bacterial rna polymerase.
Cell 2001 Mar 23;104(6):901-12
PMID: 11290327
Abstract  |  Full Article



Markovtsov, V., Mustaev, A., and Goldfarb A. (1996).
Protein-RNA interactions in the active center of transcription elongation complex. Proc. Nat'l Acad. Sci. USA, 93:3221-3226

Severinov, K., Mustaev, A., Severinova, E., Kozlov, M., Darst, S.A. and Goldfarb, A. (1995).
The beta subunit Rif-cluster I is only angstroms away from the active center of Escherichia coli RNA polymerase.
J. Biol. Chem., 270:29428-29432.

Severinov., K., , D., Severinova, E., Nikiforov, V., Landick, R., Darst, S.A., and Goldfarb, A. (1995).
Streptolydigin-resistant mutants in an evolutionarily conserved region of the beta' subunit of Escherichia coli RNA polymerase.
J. Biol. Chem., 270:23926-23929.

Nudler, E., Kashlev, M., Nikiforov, V., and Goldfarb A. (1995).
Coupling between transcription termination and RNA polymerase inchworming. Cell, 81:351-157.

Stebbins, C.E., Borukhov, S., Orlova, M., Polyakov, A., Goldfarb A., and Darst S.A. (1994).
Crystal structure of the GreA transcript cleavage factor from Escherichia coli.
Nature, 373:636-640.

PubMed Lisitings>




C.V.

Ph.D., Moscow State University, M.S., 1969; Weizmann Institute of Science, Israel, Ph.D., 1980; Max Planck Institute, Germany, Post Doc., 1980-1981; Columbia University Department of Microbiology, Assistant Professor, 1982-1991; PHRI, 1992-present.

 
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