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Biochim Biophys Acta. 2013 Feb;1829(2):187-98. doi: 10.1016/j.bbagrm.2012.11.005. Epub 2012 Nov 30.

The RNA polymerase bridge helix YFI motif in catalysis, fidelity and translocation.

Author information

1
Department of Biochemistry and Molecular Biology, Michigan State University, E. Lansing, MI 48824-1319, USA.

Abstract

The bridge α-helix in the β' subunit of RNA polymerase (RNAP) borders the active site and may have roles in catalysis and translocation. In Escherichia coli RNAP, a bulky hydrophobic segment near the N-terminal end of the bridge helix is identified (β' 772-YFI-774; the YFI motif). YFI is located at a distance from the active center and adjacent to a glycine hinge (β' 778-GARKG-782) involved in dynamic bending of the bridge helix. Remarkably, amino acid substitutions in YFI significantly alter intrinsic termination, pausing, fidelity and translocation of RNAP. F773V RNAP largely ignores the λ tR2 terminator at 200μM NTPs and is strongly reduced in λ tR2 recognition at 1μM NTPs. F773V alters RNAP pausing and backtracking and favors misincorporation. By contrast, the adjacent Y772A substitution increases fidelity and exhibits other transcriptional defects generally opposite to those of F773V. All atom molecular dynamics simulation revealed two separate functional connections emanating from YFI explaining the distinct effects of substitutions: Y772 communicates with the active site through the link domain in the β subunit, whereas F773 communicates through the fork domain in the β subunit. I774 interacts with the F-loop, which also contacts the glycine hinge of the bridge helix. These results identified negative and positive circuits coupled at YFI and employed for regulation of catalysis, elongation, termination and translocation.

PMID:
23202476
PMCID:
PMC3619131
DOI:
10.1016/j.bbagrm.2012.11.005
[Indexed for MEDLINE]
Free PMC Article

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