Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.

The actin capping protein (CP) tightly binds to the barbed end of actin filaments, thus playing a key role in actin-based lamellipodial dynamics. V-1 and CARMIL proteins directly bind to CP and inhibit the filament capping activity of CP. V-1 completely inhibits CP from interacting with the barbed e...

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Autores principales:	Shuichi Takeda, Shiho Minakata, Ryotaro Koike, Ichiro Kawahata, Akihiro Narita, Masashi Kitazawa, Motonori Ota, Tohru Yamakuni, Yuichiro Maéda, Yasushi Nitanai
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/69389e1686704efa80a266ffc4ecf1dd
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spelling	oai:doaj.org-article:69389e1686704efa80a266ffc4ecf1dd2021-12-02T19:54:46ZTwo distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.1544-91731545-788510.1371/journal.pbio.1000416https://doaj.org/article/69389e1686704efa80a266ffc4ecf1dd2010-07-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/20625546/pdf/?tool=EBIhttps://doaj.org/toc/1544-9173https://doaj.org/toc/1545-7885The actin capping protein (CP) tightly binds to the barbed end of actin filaments, thus playing a key role in actin-based lamellipodial dynamics. V-1 and CARMIL proteins directly bind to CP and inhibit the filament capping activity of CP. V-1 completely inhibits CP from interacting with the barbed end, whereas CARMIL proteins act on the barbed end-bound CP and facilitate its dissociation from the filament (called uncapping activity). Previous studies have revealed the striking functional differences between the two regulators. However, the molecular mechanisms describing how these proteins inhibit CP remains poorly understood. Here we present the crystal structures of CP complexed with V-1 and with peptides derived from the CP-binding motif of CARMIL proteins (CARMIL, CD2AP, and CKIP-1). V-1 directly interacts with the primary actin binding surface of CP, the C-terminal region of the alpha-subunit. Unexpectedly, the structures clearly revealed the conformational flexibility of CP, which can be attributed to a twisting movement between the two domains. CARMIL peptides in an extended conformation interact simultaneously with the two CP domains. In contrast to V-1, the peptides do not directly compete with the barbed end for the binding surface on CP. Biochemical assays revealed that the peptides suppress the interaction between CP and V-1, despite the two inhibitors not competing for the same binding site on CP. Furthermore, a computational analysis using the elastic network model indicates that the interaction of the peptides alters the intrinsic fluctuations of CP. Our results demonstrate that V-1 completely sequesters CP from the barbed end by simple steric hindrance. By contrast, CARMIL proteins allosterically inhibit CP, which appears to be a prerequisite for the uncapping activity. Our data suggest that CARMIL proteins down-regulate CP by affecting its conformational dynamics. This conceptually new mechanism of CP inhibition provides a structural basis for the regulation of the barbed end elongation in cells.Shuichi TakedaShiho MinakataRyotaro KoikeIchiro KawahataAkihiro NaritaMasashi KitazawaMotonori OtaTohru YamakuniYuichiro MaédaYasushi NitanaiPublic Library of Science (PLoS)articleBiology (General)QH301-705.5ENPLoS Biology, Vol 8, Iss 7, p e1000416 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Biology (General) QH301-705.5
spellingShingle	Biology (General) QH301-705.5 Shuichi Takeda Shiho Minakata Ryotaro Koike Ichiro Kawahata Akihiro Narita Masashi Kitazawa Motonori Ota Tohru Yamakuni Yuichiro Maéda Yasushi Nitanai Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
description	The actin capping protein (CP) tightly binds to the barbed end of actin filaments, thus playing a key role in actin-based lamellipodial dynamics. V-1 and CARMIL proteins directly bind to CP and inhibit the filament capping activity of CP. V-1 completely inhibits CP from interacting with the barbed end, whereas CARMIL proteins act on the barbed end-bound CP and facilitate its dissociation from the filament (called uncapping activity). Previous studies have revealed the striking functional differences between the two regulators. However, the molecular mechanisms describing how these proteins inhibit CP remains poorly understood. Here we present the crystal structures of CP complexed with V-1 and with peptides derived from the CP-binding motif of CARMIL proteins (CARMIL, CD2AP, and CKIP-1). V-1 directly interacts with the primary actin binding surface of CP, the C-terminal region of the alpha-subunit. Unexpectedly, the structures clearly revealed the conformational flexibility of CP, which can be attributed to a twisting movement between the two domains. CARMIL peptides in an extended conformation interact simultaneously with the two CP domains. In contrast to V-1, the peptides do not directly compete with the barbed end for the binding surface on CP. Biochemical assays revealed that the peptides suppress the interaction between CP and V-1, despite the two inhibitors not competing for the same binding site on CP. Furthermore, a computational analysis using the elastic network model indicates that the interaction of the peptides alters the intrinsic fluctuations of CP. Our results demonstrate that V-1 completely sequesters CP from the barbed end by simple steric hindrance. By contrast, CARMIL proteins allosterically inhibit CP, which appears to be a prerequisite for the uncapping activity. Our data suggest that CARMIL proteins down-regulate CP by affecting its conformational dynamics. This conceptually new mechanism of CP inhibition provides a structural basis for the regulation of the barbed end elongation in cells.
format	article
author	Shuichi Takeda Shiho Minakata Ryotaro Koike Ichiro Kawahata Akihiro Narita Masashi Kitazawa Motonori Ota Tohru Yamakuni Yuichiro Maéda Yasushi Nitanai
author_facet	Shuichi Takeda Shiho Minakata Ryotaro Koike Ichiro Kawahata Akihiro Narita Masashi Kitazawa Motonori Ota Tohru Yamakuni Yuichiro Maéda Yasushi Nitanai
author_sort	Shuichi Takeda
title	Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
title_short	Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
title_full	Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
title_fullStr	Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
title_full_unstemmed	Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
title_sort	two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/69389e1686704efa80a266ffc4ecf1dd
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Two distinct mechanisms for actin capping protein regulation--steric and allosteric inhibition.

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