Atomic Group "Mutagenesis" Reveals Major Groove Fine Interactions of a tRNA Synthetase with an RNA Helix
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- 作者：Penny J. Beuning ; Miriam Gulotta ; and Karin Musier-Forsyth
- 刊名：Journal of the American Chemical Society
- 出版年：1997
- 出版时间：September 10, 1997
- 年：1997
- 卷：119
- 期：36
- 页码：8397 - 8402
- 全文大小：215K
- 年卷期：v.119,no.36(September 10, 1997)
- ISSN：1520-5126
文摘
RNA discrimination by aminoacyl-tRNA synthetases involves bothmajor and minor groove interactionswith the acceptor stem domain of tRNA substrates. In the case ofclass II Escherichia coli alanyl-tRNAsynthetase(AlaRS), minor groove atomic groups in and around the unique G3:U70base pair previously have been shown to becritical for recognition. In this work, we probe the role of thefirst (1:72) base pair in discrimination by AlaRS byincorporating 26 new base pair combinations at this site. We findthat atomic groups in the wild-type G1:C72 basepair do not contribute as significantly to positive recognition byAlaRS as the minor groove elements in and aroundthe G:U base pair. Our results, however, are consistent with theimportance of major groove discrimination at thissite. In particular, substrates with a major groove carbonyloxygen presented by either a G or a U at position 72 arevery poor alanine acceptors. Comparison of inactive N1:G72duplexAla variants with activeN1:2-aminopurine72variants shows that deletion of the 6-keto oxygen and the N1-hydrogenof G72 results in a transition state stabilizationof at least 3.0 kcal/mol. This work provides an example of asystem that combines minor groove interactions at aninternal position with the high selectivity of major grooveinteractions that are possible at the end of an RNA helix.