PMID: 6164388Mar 17, 1981Paper

Early steps in the path of nascent ribonucleic acid across the surface of ribonucleic acid polymerase, determined by photoaffinity labeling

Biochemistry
L H DeRiemer, C F Meares

Abstract

The photoaffinity probes beta-(4-azidophenyl) adenosine 5'-diphosphate (N3PhppA) and beta-(4-azidophenyl) adenylyl-(3'--5')-uridine 5'-diphosphate (N3PhppApU) were used to determine the RNA polymerase subunit contacts made by the 5' ends of three nascent RNA chains. Ternary enzyme-poly[d(A-T)].oligonucleotide complexes were prepared in which the nascent oligonucleotide contained a photoaffinity label at the 5' end and a 32P radiolabel only at the 3' end. The length of the RNA was fixed at two, three, or four nucleotides. Photolysis of the ternary complexes was followed by dissociation, polyacrylamide gel electrophoresis, autoradiography, and scintillation counting. With a dinucleotide probe, the enzyme subunits labeled were beta' (71%) and sigma (21%). Photolysis of the ternary complex containing trinucleotide RNA also resulted in labeling of the beta' (64%) and sigma (35%) subunits. With a tetranucleotide, the beta' subunit was very heavily labeled (88%), and a small amount of labeling of the beta (7%) and sigma (4%) subunits was observed. The alpha subunit was not labeled with any of the probes. These results imply that a conformational change, possibly involving dissociation of the sigma subunit, occurs in the enzyme as the ...Continue Reading

References

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Citations

Jan 1, 1988·Journal of Molecular Evolution·A LazcanoJ Oró
Dec 14, 1984·Biochemical and Biophysical Research Communications·E Bateman, B H Nicholson
Dec 19, 2001·Journal of Photochemistry and Photobiology. B, Biology·P V AlekseyevT S Godovikova
Jul 1, 1983·Proceedings of the National Academy of Sciences of the United States of America·M M Hanna, C F Meares
Jan 11, 2002·Molecular and Cellular Biology·Jennifer F Kugel, James A Goodrich
Jan 1, 1985·CRC Critical Reviews in Biochemistry·A Sentenac
Jul 26, 2002·Analytical Chemistry·Zoraida P AguilarIngrid Fritsch

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