Genetic encoding of 3‐nitro‐tyrosine reveals the impacts of 14‐3‐3 nitration on client binding and dephosphorylation
14‐3‐3 proteins are central hub regulators of hundreds of phosphorylated “client” proteins. They are subject to over 60 post‐translational modifications (PTMs), yet little is known how these PTMs alter 14‐3‐3 function and its ability to regulate downstream signaling pathways. An often neglected, but...
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Published in | Protein science Vol. 32; no. 3; pp. e4574 - n/a |
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Main Authors | , , , , , |
Format | Journal Article |
Language | English |
Published |
Hoboken, USA
John Wiley & Sons, Inc
01.03.2023
Wiley Subscription Services, Inc Wiley Blackwell (John Wiley & Sons) |
Subjects | |
Online Access | Get full text |
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Summary: | 14‐3‐3 proteins are central hub regulators of hundreds of phosphorylated “client” proteins. They are subject to over 60 post‐translational modifications (PTMs), yet little is known how these PTMs alter 14‐3‐3 function and its ability to regulate downstream signaling pathways. An often neglected, but well‐documented 14‐3‐3 PTM found under physiological and immune‐stimulatory conditions is the conversion of tyrosine to 3‐nitro‐tyrosine at several Tyr sites, two of which are located at sites considered important for 14‐3‐3 function: Y130 (β‐isoform numbering) is located in the primary phospho‐client peptide‐binding groove, while Y213 is found on a secondary binding site that engages with clients for full 14‐3‐3/client complex formation and client regulation. By genetically encoding 3‐nitro‐tyrosine, we sought to understand if nitration at Y130 and Y213 effectively modulated 14‐3‐3 structure, function, and client complexation. The 1.5 Å resolution crystal structure of 14‐3‐3 nitrated at Y130 showed the nitro group altered the conformation of key residues in the primary binding site, while functional studies confirmed client proteins failed to bind this variant of 14‐3‐3. But, in contrast to other client‐binding deficient variants, it did not localize to the nucleus. The 1.9 Å resolution structure of 14‐3‐3 nitrated at Y213 revealed unusual flexibility of its C‐terminal α‐helix resulting in domain swapping, suggesting additional structural plasticity though its relevance is not clear as this nitrated form retained its ability to bind clients. Collectively, our data suggest that nitration of 14‐3‐3 will alter downstream signaling systems, and if uncontrolled could result in global dysregulation of the 14‐3‐3 interactome.
PDB Code(s): 8EQ8 and 8EQH; |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 USDOE Review Editor: John Kuriyan Funding information National Institute of General Medical Science, Grant/Award Numbers: P30 GM124169‐01, RM1‐GM144227; National Institute of Health, Grant/Award Numbers: 1S10OD020111‐01, 5R01GM1114653‐04, 5R01GM131168‐02; Oregon Health Sciences University Medical Research Foundatin; Collins Medical Trust; U.S. DOE Office of Science, Grant/Award Number: DE‐AC02‐05CH11231; Russian Science Foundation, Grant/Award Number: 19‐74‐10031 |
ISSN: | 0961-8368 1469-896X 1469-896X |
DOI: | 10.1002/pro.4574 |