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Comprehensive Characterization of (S)GTP-Binding Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition Assays
[Image: see text] Thiopurine drugs are widely used as antileukemic drugs and immunosuppressive agents, and 6-thioguanosine triphosphate ((S)GTP) is a major metabolite for these drugs. Recent studies have suggested that thiopurine drugs may exert their cytotoxic effects partly through binding of (S)G...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American
Chemical
Society
2014
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4014148/ https://www.ncbi.nlm.nih.gov/pubmed/24689502 http://dx.doi.org/10.1021/ac500588q |
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author | Xiao, Yongsheng Ji, Debin Guo, Lei Wang, Yinsheng |
author_facet | Xiao, Yongsheng Ji, Debin Guo, Lei Wang, Yinsheng |
author_sort | Xiao, Yongsheng |
collection | PubMed |
description | [Image: see text] Thiopurine drugs are widely used as antileukemic drugs and immunosuppressive agents, and 6-thioguanosine triphosphate ((S)GTP) is a major metabolite for these drugs. Recent studies have suggested that thiopurine drugs may exert their cytotoxic effects partly through binding of (S)GTP to a GTP-binding protein, Rac1. However, it remains unclear whether (S)GTP can also bind to other cellular proteins. Here, we introduced an orthogonal approach, encompassing nucleotide-affinity profiling and nucleotide-binding competition assays, to characterize comprehensively (S)GTP-binding proteins along with the specific binding sites from the entire human proteome. With the simultaneous use of (S)GTP and GTP affinity probes, we identified 165 (S)GTP-binding proteins that are involved in several different biological processes. We also examined the binding selectivities of these proteins toward (S)GTP and GTP, which allowed for the revelation of the relative binding affinities of the two nucleotides toward the nucleotide-binding motif sequence of proteins. Our results suggest that (S)GTP mainly targets GTPases, with strong binding affinities observed for multiple heterotrimeric G proteins. We also demonstrated that (S)GTP binds to several cyclin-dependent kinases (CDKs), which may perturb the CDK-mediated phosphorylation and cell cycle progression. Together, this represents the first comprehensive characterization of (S)GTP-binding property for the entire human proteome. We reason that a similar strategy can be generally employed for the future characterization of the interaction of other modified nucleotides with the global proteome. |
format | Online Article Text |
id | pubmed-4014148 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | American
Chemical
Society |
record_format | MEDLINE/PubMed |
spelling | pubmed-40141482015-04-01 Comprehensive Characterization of (S)GTP-Binding Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition Assays Xiao, Yongsheng Ji, Debin Guo, Lei Wang, Yinsheng Anal Chem [Image: see text] Thiopurine drugs are widely used as antileukemic drugs and immunosuppressive agents, and 6-thioguanosine triphosphate ((S)GTP) is a major metabolite for these drugs. Recent studies have suggested that thiopurine drugs may exert their cytotoxic effects partly through binding of (S)GTP to a GTP-binding protein, Rac1. However, it remains unclear whether (S)GTP can also bind to other cellular proteins. Here, we introduced an orthogonal approach, encompassing nucleotide-affinity profiling and nucleotide-binding competition assays, to characterize comprehensively (S)GTP-binding proteins along with the specific binding sites from the entire human proteome. With the simultaneous use of (S)GTP and GTP affinity probes, we identified 165 (S)GTP-binding proteins that are involved in several different biological processes. We also examined the binding selectivities of these proteins toward (S)GTP and GTP, which allowed for the revelation of the relative binding affinities of the two nucleotides toward the nucleotide-binding motif sequence of proteins. Our results suggest that (S)GTP mainly targets GTPases, with strong binding affinities observed for multiple heterotrimeric G proteins. We also demonstrated that (S)GTP binds to several cyclin-dependent kinases (CDKs), which may perturb the CDK-mediated phosphorylation and cell cycle progression. Together, this represents the first comprehensive characterization of (S)GTP-binding property for the entire human proteome. We reason that a similar strategy can be generally employed for the future characterization of the interaction of other modified nucleotides with the global proteome. American Chemical Society 2014-04-01 2014-05-06 /pmc/articles/PMC4014148/ /pubmed/24689502 http://dx.doi.org/10.1021/ac500588q Text en Copyright © 2014 American Chemical Society |
spellingShingle | Xiao, Yongsheng Ji, Debin Guo, Lei Wang, Yinsheng Comprehensive Characterization of (S)GTP-Binding Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition Assays |
title | Comprehensive Characterization of (S)GTP-Binding
Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition
Assays |
title_full | Comprehensive Characterization of (S)GTP-Binding
Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition
Assays |
title_fullStr | Comprehensive Characterization of (S)GTP-Binding
Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition
Assays |
title_full_unstemmed | Comprehensive Characterization of (S)GTP-Binding
Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition
Assays |
title_short | Comprehensive Characterization of (S)GTP-Binding
Proteins by Orthogonal Quantitative (S)GTP-Affinity Profiling and (S)GTP/GTP Competition
Assays |
title_sort | comprehensive characterization of (s)gtp-binding
proteins by orthogonal quantitative (s)gtp-affinity profiling and (s)gtp/gtp competition
assays |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4014148/ https://www.ncbi.nlm.nih.gov/pubmed/24689502 http://dx.doi.org/10.1021/ac500588q |
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