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Title: Structure and two-metal mechanism of fungal tRNA ligase

Journal Article · · Nucleic Acids Research
DOI:https://doi.org/10.1093/nar/gky1275· OSTI ID:1488757
 [1];  [1];  [2];  [1]
  1. Molecular Biology Program, Sloan-Kettering Institute, New York, NY 10065, USA
  2. Structural Biology Program, Sloan-Kettering Institute, New York, NY 10065, USA
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Fungal tRNA ligase (Trl1) is an essential enzyme that repairs RNA breaks with 2',3;-cyclic-PO4 and 5'-OH ends inflicted during tRNA splicing and non-canonical mRNA splicing in the fungal unfolded protein response. Trl1 is composed of C-terminal cyclic phosphodiesterase (CPD) and central GTP-dependent polynucleotide kinase (KIN) domains that heal the broken ends to generate the 3'-OH,2'-PO4 and 5'-PO4 termini required for sealing by an N-terminal ATP-dependent ligase domain (LIG). Here we report crystal structures of the Trl1-LIG domain from Chaetomium thermophilum at two discrete steps along the reaction pathway: the covalent LIG-(lysyl-Nζ)–AMP•Mn2+ intermediate and a LIG•ATP•(Mn2+)2 Michaelis complex. The structures highlight a two-metal mechanism whereby a penta-hydrated metal complex stabilizes the transition state of the ATP α phosphate and a second metal bridges the β and γ phosphates to help orient the pyrophosphate leaving group. A LIG-bound sulfate anion is a plausible mimetic of the essential RNA terminal 2'-PO4. Trl1-LIG has a distinctive C-terminal domain that instates fungal Trl1 as the founder of an Rnl6 clade of ATP-dependent RNA ligase. We discuss how the Trl1-LIG structure rationalizes the large body of in vivo structure–function data for Saccharomyces cerevisiae Trl1.

Research Organization:
Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Organization:
USDOE; National Institutes of Health (NIH)
Grant/Contract Number:
AC02-06CH11357; R35-GM126945
OSTI ID:
1488757
Alternate ID(s):
OSTI ID: 1502195
Journal Information:
Nucleic Acids Research, Journal Name: Nucleic Acids Research Vol. 47 Journal Issue: 3; ISSN 0305-1048
Publisher:
Oxford University PressCopyright Statement
Country of Publication:
United Kingdom
Language:
English
Citation Metrics:
Cited by: 9 works
Citation information provided by
Web of Science

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