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Novel activity of a yeast ligase deletion polypeptide. Evidence for GTP-dependent tRNA splicing.

Westaway, Shawn K. and Belford, Heather G. and Apostol, Barbara L. and Abelson, John and Greer, Chris L. (1993) Novel activity of a yeast ligase deletion polypeptide. Evidence for GTP-dependent tRNA splicing. Journal of Biological Chemistry, 268 (4). pp. 2435-2443. ISSN 0021-9258. https://resolver.caltech.edu/CaltechAUTHORS:20140312-113308036

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Abstract

Yeast tRNA ligase possesses multiple activities which are required for the joining of tRNA halves during the tRNA splicing process: cyclic phosphodiesterase, kinase, adenylylate synthetase, and ligase. A deletion polypeptide of a dihydrofolate reductase-ligase fusion protein, designated DAC, was previously shown to join tRNA halves although ATP-dependent kinase activity was not measurable in the assay used. We describe here a characterization of the mechanism of joining used by DAC and the structure of the tRNA product. DAC produces a joined tRNA and a splice junction with a structure identical to that produced by DAKC, the full-length dihydrofolate reductase-ligase fusion. Furthermore, DAC can use GTP as the sole cofactor in the joining reaction, in contrast to DAKC, which can only complete splicing in the presence of ATP. Both enzymes exhibit GTP-dependent kinase activity at 100-fold greater efficiency than with ATP. These results suggest that a potential function for the center domain of tRNA ligase (missing in DAC) is to provide structural integrity and aid in substrate interactions and specificity. They also support the hypothesis that ligase may prefer to use two different cofactors during tRNA splicing.


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http://www.jbc.org/content/268/4/2435.abstract?sid=3a12edae-e3a1-4925-ac55-5b83c6788898PublisherArticle
Additional Information:© 1993 The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, July 1, 1992. This work was supported by Grants DMB-891393 from the National Science Foundation and GM-35955 from the National Institutes of Health (to C. L. G.) and by American Cancer Society Grant MV-318K (to J. A,). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. We thank Reinhard Rauhut for generously providing purified yeast tRNA splicing endonuclease and for critical review of the manuscript. We are grateful to Craig Peebles, Eric Phizicky, Rosalind Haselbeck, Mark Berlin, and Daniel Herschlag for helpful discussions and comments.
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Funding AgencyGrant Number
NSFDMB-891393
NIHGM-35955
American Cancer SocietyMV-318K
Issue or Number:4
Record Number:CaltechAUTHORS:20140312-113308036
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20140312-113308036
Official Citation:Westaway, S. K., Belford, H. G., Apostol, B. L., Abelson, J., & Greer, C. L. (1993). Novel activity of a yeast ligase deletion polypeptide. Evidence for GTP-dependent tRNA splicing. Journal of Biological Chemistry, 268(4), 2435-2443.
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:44279
Collection:CaltechAUTHORS
Deposited By: Jason Perez
Deposited On:12 Mar 2014 20:10
Last Modified:03 Oct 2019 06:16

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