气候变化领域集成服务门户(CCPortal): Trigger loop of RNA polymerase is a positional not acid-base; catalyst for both transcription and proofreading

CCPortal

DOI	10.1073/pnas.1702383114
	Trigger loop of RNA polymerase is a positional not acid-base; catalyst for both transcription and proofreading
	Mishanina T.V.; Palo M.Z.; Nayak D.; Mooney R.A.; Landick R.
发表日期	2017
ISSN	0027-8424
起始页码	E5103
结束页码	E5112
卷号	114 期号:26
英文摘要	The active site of multisubunit RNA polymerases (RNAPs) is highly conserved from humans to bacteria. This single site catalyzes both nucleotide addition required for RNA transcript synthesis and excision of incorrect nucleotides after misincorporation as a proofreading mechanism. Phosphoryl transfer and proofreading hydrolysis are controlled in part by a dynamic RNAP component called the trigger loop (TL), which cycles between an unfolded loop and an α-helical hairpin [trigger helices (TH)] required for rapid nucleotide addition. The precise roles of the TL/TH in RNA synthesis and hydrolysis remain unclear. An invariant histidine residue has been proposed to function in the TH form as a general acid in RNA synthesis and as a general base in RNA hydrolysis. The effects of conservative, nonionizable substitutions of the TL histidine (or a neighboring TL arginine conserved in bacteria) have not yet been rigorously tested. Here, we report that glutamine substitutions of these residues, which preserve polar interactions but are incapable of acid-base chemistry, had little effect on either phosphoryl transfer or proofreading hydrolysis by Escherichia coli RNAP. The TL substitutions did, however, affect the backtracking of RNAP necessary for proofreading and potentially the reactivity of the backtracked nucleotide. We describe a unifying model for the function of the RNAP TL, which reconciles available data and our results for representative RNAPs. This model explains diverse effects of the TL basic residues on catalysis through their effects on positioning reactants for phosphoryl transfer and easing barriers to transcript backtracking, rather than as acid-base catalysts.
英文关键词	Acid-base catalysis; Proofreading; RNA hydrolysis; Transcription; Trigger loop
语种	英语
scopus关键词	acid; base; glutamine; molecular scaffold; RNA polymerase; DNA directed RNA polymerase; Escherichia coli protein; RNA; Article; catalyst; enzyme active site; enzyme reconstitution; Escherichia coli; genetic transcription; hydrolysis; nonhuman; priority journal; substitution reaction; amino acid substitution; biosynthesis; chemical model; chemistry; conformation; enzymology; genetics; metabolism; missense mutation; Amino Acid Substitution; DNA-Directed RNA Polymerases; Escherichia coli; Escherichia coli Proteins; Models, Chemical; Mutation, Missense; Nucleic Acid Conformation; RNA
来源期刊	Proceedings of the National Academy of Sciences of the United States of America
文献类型	期刊论文
条目标识符	http://gcip.llas.ac.cn/handle/2XKMVOVA/160621
作者单位	Mishanina, T.V., Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, United States, Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC 29425, United States; Palo, M.Z., Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, United States; Nayak, D., Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, United States; Mooney, R.A., Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, United States; Landick, R., Department of Biochemistry, University of Wisconsin-Madison, Madison, WI 53706, United States
推荐引用方式 GB/T 7714	Mishanina T.V.,Palo M.Z.,Nayak D.,et al. Trigger loop of RNA polymerase is a positional not acid-base; catalyst for both transcription and proofreading[J],2017,114(26).
APA	Mishanina T.V.,Palo M.Z.,Nayak D.,Mooney R.A.,&Landick R..(2017).Trigger loop of RNA polymerase is a positional not acid-base; catalyst for both transcription and proofreading.Proceedings of the National Academy of Sciences of the United States of America,114(26).
MLA	Mishanina T.V.,et al."Trigger loop of RNA polymerase is a positional not acid-base; catalyst for both transcription and proofreading".Proceedings of the National Academy of Sciences of the United States of America 114.26(2017).