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A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity

A protoberberine derivative library was used to search for selective inhibitors against kinases of the mitogen-activated protein kinase (MAPK) cascades in mammalian cells. Among kinases in mammalian MAPK pathways, we identified a compound (HWY336) that selectively inhibits kinase activity of mitogen...

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Autores principales: Kim, Namil, Park, Jeongyeon, Gadhe, Changdev G., Cho, Seung Joo, Oh, Youngjin, Kim, Donghyun, Song, Kiwon
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3997336/
https://www.ncbi.nlm.nih.gov/pubmed/24759688
http://dx.doi.org/10.1371/journal.pone.0091037
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author Kim, Namil
Park, Jeongyeon
Gadhe, Changdev G.
Cho, Seung Joo
Oh, Youngjin
Kim, Donghyun
Song, Kiwon
author_facet Kim, Namil
Park, Jeongyeon
Gadhe, Changdev G.
Cho, Seung Joo
Oh, Youngjin
Kim, Donghyun
Song, Kiwon
author_sort Kim, Namil
collection PubMed
description A protoberberine derivative library was used to search for selective inhibitors against kinases of the mitogen-activated protein kinase (MAPK) cascades in mammalian cells. Among kinases in mammalian MAPK pathways, we identified a compound (HWY336) that selectively inhibits kinase activity of mitogen-activated protein kinase kinase 4 and 7 (MKK4 and MKK7). The IC(50) of HWY336 was 6 µM for MKK4 and 10 µM for MKK7 in vitro. HWY336 bound to both kinases reversibly via noncovalent interactions, and inhibited their activity by interfering with access of a protein substrate to its binding site. The binding affinity of HWY336 to MKK4 was measured by surface plasmon resonance to determine a dissociation constant (K(d)) of 3.2 µM. When mammalian cells were treated with HWY336, MKK4 and MKK7 were selectively inhibited, resulting in inhibition of c-Jun NH(2)-terminal protein kinases in vivo. The structural model of HWY336 bound to either MKK4 or MKK7 predicted that HWY336 was docked to the activation loop, which is adjacent to the substrate binding site. This model suggested the importance of the activation loop of MKKs in HWY336 selectivity. We verified this model by mutating three critical residues within this loop of MKK4 to the corresponding residues in MKK3. The mutant MKK4 displayed similar kinase activity as wild-type kinase, but its activity was not inhibited by HWY336 compared to wild-type MKK4. We propose that the specific association of HWY336 to the activation loop of MKK4/MKK7 is responsible for its selective inhibition.
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spelling pubmed-39973362014-04-29 A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity Kim, Namil Park, Jeongyeon Gadhe, Changdev G. Cho, Seung Joo Oh, Youngjin Kim, Donghyun Song, Kiwon PLoS One Research Article A protoberberine derivative library was used to search for selective inhibitors against kinases of the mitogen-activated protein kinase (MAPK) cascades in mammalian cells. Among kinases in mammalian MAPK pathways, we identified a compound (HWY336) that selectively inhibits kinase activity of mitogen-activated protein kinase kinase 4 and 7 (MKK4 and MKK7). The IC(50) of HWY336 was 6 µM for MKK4 and 10 µM for MKK7 in vitro. HWY336 bound to both kinases reversibly via noncovalent interactions, and inhibited their activity by interfering with access of a protein substrate to its binding site. The binding affinity of HWY336 to MKK4 was measured by surface plasmon resonance to determine a dissociation constant (K(d)) of 3.2 µM. When mammalian cells were treated with HWY336, MKK4 and MKK7 were selectively inhibited, resulting in inhibition of c-Jun NH(2)-terminal protein kinases in vivo. The structural model of HWY336 bound to either MKK4 or MKK7 predicted that HWY336 was docked to the activation loop, which is adjacent to the substrate binding site. This model suggested the importance of the activation loop of MKKs in HWY336 selectivity. We verified this model by mutating three critical residues within this loop of MKK4 to the corresponding residues in MKK3. The mutant MKK4 displayed similar kinase activity as wild-type kinase, but its activity was not inhibited by HWY336 compared to wild-type MKK4. We propose that the specific association of HWY336 to the activation loop of MKK4/MKK7 is responsible for its selective inhibition. Public Library of Science 2014-04-23 /pmc/articles/PMC3997336/ /pubmed/24759688 http://dx.doi.org/10.1371/journal.pone.0091037 Text en © 2014 Kim et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Kim, Namil
Park, Jeongyeon
Gadhe, Changdev G.
Cho, Seung Joo
Oh, Youngjin
Kim, Donghyun
Song, Kiwon
A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title_full A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title_fullStr A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title_full_unstemmed A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title_short A Protoberberine Derivative HWY336 Selectively Inhibits MKK4 and MKK7 in Mammalian Cells: The Importance of Activation Loop on Selectivity
title_sort protoberberine derivative hwy336 selectively inhibits mkk4 and mkk7 in mammalian cells: the importance of activation loop on selectivity
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3997336/
https://www.ncbi.nlm.nih.gov/pubmed/24759688
http://dx.doi.org/10.1371/journal.pone.0091037
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