Fluorescence mapping of the open complex of yeast mitochondrial RNA polymerase.

The Journal of Biological Chemistry
Guo-Qing TangS S Patel

Abstract

The mitochondrial RNA polymerase (mtRNAP) of Saccharomyces cerevisiae, consisting of a complex of Rpo41 and Mtf1, is homologous to the phage single polypeptide T7/T3 RNA polymerases. The yeast mtRNAP recognizes a conserved nonanucleotide sequence to initiate specific transcription. In this work, we have defined the region of the nonanucleotide that is melted by the mtRNAP using 2-aminopurine (2AP) fluorescence that is sensitive to changes in base stacking interactions. We show that mtRNAP spontaneously melts the promoter from -4 to +2 forming a bubble around the transcription start site at +1. The location and size of the DNA bubble in this open complex of the mtRNAP closely resembles that of the T7 RNA polymerase. We show that DNA melting requires the simultaneous presence of Rpo41 and Mtf1. Adding the initiating nucleotide ATP does not expand the size of the initially melted DNA, but the initiating nucleotide differentially affects base stacking interactions at -1 and -2. Thus, the promoter structure upstream of the transcription start site is slightly rearranged during early initiation from its structure in the pre-initiation stage. Unlike on the duplex promoter, Rpo41 alone was able to form a competent open complex on a pre...Continue Reading

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Citations

Dec 17, 2009·The Journal of Biological Chemistry·Swaroopa Paratkar, Smita S Patel
Nov 19, 2009·The Journal of Biological Chemistry·Maria SavkinaMichael Anikin
Mar 31, 2010·The Journal of Biological Chemistry·Maria F LodeiroCraig E Cameron
Apr 2, 2011·The Journal of Biological Chemistry·Swaroopa ParatkarSmita S Patel
Sep 14, 2011·The Journal of Biological Chemistry·Guo-Qing TangSmita S Patel
Sep 14, 2011·Nucleic Acids Research·Hajin KimTaekjip Ha
Feb 23, 2012·Biochimica Et Biophysica Acta·Aishwarya P Deshpande, Smita S Patel
Feb 4, 2011·Chemical Society Reviews·Nan Dai, Eric T Kool
Aug 29, 2020·Nature Communications·Byeong-Kwon SohnHajin Kim
Nov 1, 2020·The Journal of Biological Chemistry·Urmimala BasuSmita S Patel
Jan 18, 2021·The Journal of Biological Chemistry·Urmimala BasuSmita S Patel

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