Dissertation/Thesis Abstract

Characterization of a tryptophanyl -tRNA synthetase urzyme and its intramolecular complementation establishes a model to study the catalytic mechanism and evolution of the contemporary class I aminoacyl -tRNA synthetases
by Pham, Yen Bao, Ph.D., The University of North Carolina at Chapel Hill, 2010, 106; 3409957
Abstract (Summary)

Aminoacyl-tRNA synthetases (AaRSs) comprise a crucial group of enzymes catalyzing a two-step reaction to activate amino acids and acylate their cognate tRNAs. It is widely known that highly conserved sequences and motifs divide the synthetases evenly into two distinct classes of ten members each. However, the questions of how they might have emerged and evolved and what a reasonable candidate for the ancestral enzyme would be like still remain unsolved. Answering these would not only shed light on the aaRSs evolution but could also help unravel the critical point when the extremely high kinetic barrier of amino acid activation was bypassed, allowing the synthesis of primitive peptide chains.

The research object of the studies is a tryptophanyl-tRNA synthetase (TrpRS) Urzyme (Ur-: prefix means primitive, original) which would fit the description of a potential candidate for the ancestral class I aaRS. The TrpRS Urzyme is designed to retain class I signature sequences that comprise the ATP-binding Rossmann fold. Interestingly, it has similar affinity for ATP compared to the full length wild type enzyme but surprisingly low affinity for the cognate amino acid substrate, tryptophan. This might be explained by absence of the connecting peptide 1 (CP1), an insertion that wraps around the tryptophan binding site in the contemporary enzyme and occurs at the same location as the editing domain in four other class I aaRSs (MetRS, LeuRS, IleRS, ValRS). An in cis modular complementation experiment might point out a potential role for CP1 in configuring a confirmation that has high affinity for tryptophan, functionally similar to a primitive proofreading function. Furthermore, an Urzyme containing the C-terminal domain (CTD) in cis has been constructed to further explore CTD’s role in tryptophan activation and tRNATrp acylation.

The active TrpRS Urzyme establishment confirms an essential prediction of, and is therefore, the first experiment to provide evidential support for the unique Rodin-Ohno hypothesis about the earlier existence of a sense-antisense coding system for both aaRS classes. The successful design of the TrpRS Urzyme has also provided a precedent that has led to the construction of class I and class II aaRS Urzymes of comparable lengths.

Indexing (document details)
Advisor: Carter, Charles W.
Commitee: Fried, Howard M., Kuhlman, Brian, Sancar, Aziz, Wolfenden, Richard
School: The University of North Carolina at Chapel Hill
Department: Biochemistry & Biophysics
School Location: United States -- North Carolina
Source: DAI-B 71/08, Dissertation Abstracts International
Subjects: Molecular biology, Biochemistry
Keywords: Aminoacyl-tRNA synthetases, Catalytic, Tryptophanyl-tRNA synthetase, Urzyme
Publication Number: 3409957
ISBN: 978-1-124-07855-7
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