C-Terminal Zinc-Containing Peptide Required for RNA Recognition by a Class I tRNA Synthetase

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• 作者：Elizabeth Glasfeld ; James A. Landro ; and Paul Schimmel
• 刊名：Biochemistry
• 出版年：1996
• 出版时间：April 2, 1996
• 年：1996
• 卷：35
• 期：13
• 页码：4139 - 4145
• 全文大小：362K
• 年卷期：v.35,no.13(April 2, 1996)
• ISSN：1520-4995

文摘

Escherichia coli isoleucyl-tRNA synthetase is one offive closely related class I tRNAsynthetases. The active site of the 939 amino acid polypeptide isin an N-terminal domain which containsan insertion believed essential for interactions with the tRNA acceptorhelix. The enzyme was shownpreviously to contain an essential (for function in vivo)zinc bound to a Cys₄ cluster at the C-terminalendof the polypeptide. The specific function of this zinc has beenunknown. We show here that aminoacylationactivity can be reconstituted in vitro by combining a 53amino acid zinc-containing C-terminal peptidewith a protein consisting of the remaining 886 amino acids.Reconstitution of aminoacylation is zinc-dependent. In contrast, the zinc-containing peptide is dispensablefor synthesis of isoleucyl adenylate.Affinity coelectrophoresis showed that the 53 amino acidC-terminal peptide is required specifically fortRNA binding. We propose that the zinc-containing peptide curlsback to the active site to make contactwith the acceptor helix of bound tRNA, but not with isoleucine or ATP.It is the first example of azinc-containing peptide in a class I tRNA synthetase that is essentialfor tRNA binding interactions. Thedesign of this enzyme may be part of a more general scheme for class ItRNA synthetases to acquireacceptor helix binding elements during the development of the geneticcode.