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The folding capacity of the mature domain of the dual-targeted plant tRNA nucleotidyltransferase influences organelle selection.

Author(s): Leibovitch M, Bublak D, Hanic-Joyce PJ, Tillmann B, Flinner N, Amsel D, Scharf KD, Mirus O, Joyce PB, Schleiff E

Biochem J. 2013 Aug 01;453(3):401-12 Authors: Leibovitch M, Bublak D, Hanic-Joyce PJ, Tillmann B, Flinner N, Amsel D, Scharf KD, Mirus O, Joyce PB, Schleiff E

Article GUID: 23713568
In vitro studies of disease-linked variants of human tRNA nucleotidyltransferase reveal decreased thermal stability and altered catalytic activity.

Author(s): Leibovitch M, Hanic-Joyce PJ, Joyce PBM

Biochim Biophys Acta Proteins Proteom. 2018 Apr;1866(4):527-540 Authors: Leibovitch M, Hanic-Joyce PJ, Joyce PBM

Article GUID: 29454993
Analysis of the pathogenic I326T variant of human tRNA nucleotidyltransferase reveals reduced catalytic activity and thermal stability in vitro linked to a conformational change.

Author(s): Leibovitch M, Reid NE, Victoria J, Hanic-Joyce PJ, Joyce PBM

Biochim Biophys Acta Proteins Proteom. 2019 Jun;1867(6):616-626 Authors: Leibovitch M, Reid NE, Victoria J, Hanic-Joyce PJ, Joyce PBM

Article GUID: 30959222
Title:Analysis of the pathogenic I326T variant of human tRNA nucleotidyltransferase reveals reduced catalytic activity and thermal stability in vitro linked to a conformational change.
Authors:Leibovitch M,  Reid NE,  Victoria J,  Hanic-Joyce PJ,  Joyce PBM
Link:https://www.ncbi.nlm.nih.gov/pubmed/30959222?dopt=Abstract
Category:Biochim Biophys Acta Proteins Proteom
PMID:30959222
Dept Affiliation: CHEMBIOCHEM
1 Department of Chemistry and Biochemistry and Centre for Structural and Functional Genomics, Concordia University, 7141 Sherbrooke St. W., Montréal H4B 1R6, Québec, Canada.
2 Department of Chemistry and Biochemistry and Centre for Structural and Functional Genomics, Concordia University, 7141 Sherbrooke St. W., Montréal H4B 1R6, Québec, Canada. Electronic address: paul.joyce@concordia.ca.

Description:

Analysis of the pathogenic I326T variant of human tRNA nucleotidyltransferase reveals reduced catalytic activity and thermal stability in vitro linked to a conformational change.

Biochim Biophys Acta Proteins Proteom. 2019 Jun;1867(6):616-626

Authors: Leibovitch M, Reid NE, Victoria J, Hanic-Joyce PJ, Joyce PBM

Abstract

The I326T mutation in the TRNT1 gene encoding human tRNA nucleotidyltransferase (tRNA-NT) is linked to a relatively mild form of SIFD. Previous work indicated that the I326T variant was unable to incorporate AMP into tRNAs in vitro, however, expression of the mutant allele from a strong heterologous promoter supported in vivo CCA addition to both cytosolic and mitochondrial tRNAs in a yeast strain lacking tRNA-NT. To address this discrepancy, we determined the biochemical and biophysical characteristics of the I326T variant enzyme and the related variant, I326A. Our in vitro analysis revealed that the I326T substitution decreases the thermal stability of the enzyme and causes a ten-fold reduction in enzyme activity. We propose that the structural changes in the I326T variant that lead to these altered parameters result from a rearrangement of helices within the body domain of the protein which can be probed by the inability of the monomeric enzyme to form a covalent dimer in vitro mediated by C373. In addition, we confirm that the effects of the I326T or I326A substitutions are relatively mild in vivo by demonstrating that the mutant alleles support both mitochondrial and cytosolic CCA-addition in yeast.

PMID: 30959222 [PubMed - in process]