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Modifications Modulate Anticodon Loop Dynamics and Codon Recognition in E. coli tRNA Arg1,2
Author(s) -
Sarachan Kathryn,
Cantara William,
Spears Jessica,
DeMirci Hasan,
Murphy Frank,
Ranganathan Srivathsan,
Chen Alan,
Agris Paul
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.711.21
Subject(s) - transfer rna , wobble base pair , ribosome , chemistry , biochemistry , stereochemistry , biology , genetics , rna , gene
In E. coli , three of six arginine codons (CGU, CGC and CGA) are decoded by only two tRNA Arg isoacceptors. The anticodon‐stem loop (ASL) domains of tRNA Arg1 and tRNA Arg2 differ only at position 32: tRNA Arg1 is posttranscriptionally modified to contain a 2‐thiocytidine (s 2 C 32 ). Both isoacceptors also contain naturally‐occurring inosine (I 34 ) and 2‐methyladenosine (m 2 A 37 ) modifications. To investigate the functional roles of these modifications, six ASL constructs differing in modification profile were analyzed using binding studies and structural and computational methods. Ribosome filter binding assays showed that while I 34 facilitates wobble codon binding, both s 2 C 32 and m 2 A 37 modulate that effect by negating binding to the rare CGA codon. When a non‐naturally occurring s 2 C 32 was introduced for C 32 in an unrelated S. cerevisiae tRNA ASL Leu construct also containing I 34 , the same functional restriction of CGA binding was observed. The NMR and x‐ray crystallographic structures of the ASLs showed only a minor anticodon loop perturbation upon inclusion of s 2 C 32 . The mechanism of its effect on codon binding was then investigated by molecular dynamics simulations, which suggested that both s 2 C 32 and m 2 A 37 afford the ASL greater flexibility and conformational accessibility and explained their ability to adopt one structure free in solution and two others when bound to the cognate arginyl‐tRNA synthetase or to codons on the ribosome.