Journal
NUCLEIC ACIDS RESEARCH
Volume 48, Issue 15, Pages 8740-8754Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkaa569
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Funding
- National Institutes of Health (NIH) [P01 HL029582, P01 HL076491]
- National Heart, Lung, and Blood Institute [R01 HL128300, R01 HL128268]
- National Institute of General Medical Sciences [R01 GM086430]
- National Institute of Diabetes and Digestive and Kidney Diseases [R01 DK124203, R01 DK123236]
- Lerner Research Institute, Cleveland Clinic
- National Institutes of Health [S10 OD023436]
- NIH [P01 HL029582]
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In mammalian cells, eight cytoplasmic aminoacyltRNA synthetases (AARS), and three non-synthetase proteins, reside in a large multi-tRNA synthetase complex (MSC). AARSs have critical roles in interpretation of the genetic code during protein synthesis, and in non-canonical functions unrelated to translation. Nonetheless, the structure and function of the MSC remain unclear. Partial or complete crystal structures of all MSC constituents have been reported; however, the structure of the holo-MSC has not been resolved. We have taken advantage of cross-linking mass spectrometry (XL-MS) and molecular docking to interrogate the three-dimensional architecture of the MSC in human HEK293T cells. The XL-MS approach uniquely provides structural information on flexibly appended domains, characteristic of nearly all MSC constituents. Using the MS-cleavable cross-linker, disuccinimidyl sulfoxide, inter-protein cross-links spanning all MSC constituents were observed, including cross-links between eight protein pairs not previously known to interact. Intra-protein cross-links defined new structural relationships between domains in several constituents. Unexpectedly, an asymmetric AARS distribution was observed featuring a clustering of tRNA anti-codon binding domains on one MSC face. Possibly, the non-uniform localization improves efficiency of delivery of charged tRNA's to an interacting ribosome during translation. In summary, we show a highly compact, 3D structural model of the human holo-MSC.
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