Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.

Hepatitis C virus (HCV) affects nearly 200 million people worldwide and is a leading factor for serious chronic liver diseases. For replicating HCV genome, the membrane-associated replication machinery needs to be formed by both HCV non-structural proteins including NS5A and human host factors. Rece...

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Autores principales:	Garvita Gupta, Haina Qin, Jianxing Song
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/2dfcaf8d6ead4e62adcc1a8645a9dcd9
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spelling	oai:doaj.org-article:2dfcaf8d6ead4e62adcc1a8645a9dcd92021-11-18T07:15:33ZIntrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.1932-620310.1371/journal.pone.0039261https://doaj.org/article/2dfcaf8d6ead4e62adcc1a8645a9dcd92012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22720086/pdf/?tool=EBIhttps://doaj.org/toc/1932-6203Hepatitis C virus (HCV) affects nearly 200 million people worldwide and is a leading factor for serious chronic liver diseases. For replicating HCV genome, the membrane-associated replication machinery needs to be formed by both HCV non-structural proteins including NS5A and human host factors. Recently NS5A has been identified to bind ER-anchored human VAP proteins and consequently this interaction may serve as a novel target for design of anti-HCV drugs. So far no biophysical characterization of this interaction has been reported. Here, we dissected the 243-residue VAPB into 4 and 447-residue NS5A into 10 fragments, followed by CD and NMR characterization of their structural properties. Subsequently, binding interactions between these fragments have been extensively assessed by NMR HSQC titration which is very powerful in detecting even very weak binding. The studies lead to three important findings: 1). a "fuzzy complex" is formed between the intrinsically-unstructured third domain (D3) of NS5A and the well-structured MSP domain of VAPB, with an average dissociation constant (Kd) of ~5 µM. 2). The binding-important residues on both NS5A-D3 and VAPB-MSP have been successfully mapped out, which provided experimental constraints for constructing the complex structure. In the complex, unstructured D3 binds to three surface pockets on one side of the MSP structure. Interestingly, two ALS-causing mutations T46I and P56S are also located on the D3-MSP interface. Moreover, NS5A-D3, FFAT-containing proteins and EphA4 appear to have overlapped binding interfaces on the MSP domain. 3). NS5A-D3 has been experimentally confirmed to competes with EphA4 in binding to the MSP domain, and T46I mutation of MSP dramatically abolishes its binding ability to D3. Our study not only provides essential foundation for further deciphering structure and function of the HCV replication machinery, but may also shed light on rationalizing a recent observation that a chronic HCV patient surprisingly developed ALS-like syndrome.Garvita GuptaHaina QinJianxing SongPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 6, p e39261 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Garvita Gupta Haina Qin Jianxing Song Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
description	Hepatitis C virus (HCV) affects nearly 200 million people worldwide and is a leading factor for serious chronic liver diseases. For replicating HCV genome, the membrane-associated replication machinery needs to be formed by both HCV non-structural proteins including NS5A and human host factors. Recently NS5A has been identified to bind ER-anchored human VAP proteins and consequently this interaction may serve as a novel target for design of anti-HCV drugs. So far no biophysical characterization of this interaction has been reported. Here, we dissected the 243-residue VAPB into 4 and 447-residue NS5A into 10 fragments, followed by CD and NMR characterization of their structural properties. Subsequently, binding interactions between these fragments have been extensively assessed by NMR HSQC titration which is very powerful in detecting even very weak binding. The studies lead to three important findings: 1). a "fuzzy complex" is formed between the intrinsically-unstructured third domain (D3) of NS5A and the well-structured MSP domain of VAPB, with an average dissociation constant (Kd) of ~5 µM. 2). The binding-important residues on both NS5A-D3 and VAPB-MSP have been successfully mapped out, which provided experimental constraints for constructing the complex structure. In the complex, unstructured D3 binds to three surface pockets on one side of the MSP structure. Interestingly, two ALS-causing mutations T46I and P56S are also located on the D3-MSP interface. Moreover, NS5A-D3, FFAT-containing proteins and EphA4 appear to have overlapped binding interfaces on the MSP domain. 3). NS5A-D3 has been experimentally confirmed to competes with EphA4 in binding to the MSP domain, and T46I mutation of MSP dramatically abolishes its binding ability to D3. Our study not only provides essential foundation for further deciphering structure and function of the HCV replication machinery, but may also shed light on rationalizing a recent observation that a chronic HCV patient surprisingly developed ALS-like syndrome.
format	article
author	Garvita Gupta Haina Qin Jianxing Song
author_facet	Garvita Gupta Haina Qin Jianxing Song
author_sort	Garvita Gupta
title	Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
title_short	Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
title_full	Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
title_fullStr	Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
title_full_unstemmed	Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.
title_sort	intrinsically unstructured domain 3 of hepatitis c virus ns5a forms a "fuzzy complex" with vapb-msp domain which carries als-causing mutations.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/2dfcaf8d6ead4e62adcc1a8645a9dcd9
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Intrinsically unstructured domain 3 of hepatitis C Virus NS5A forms a "fuzzy complex" with VAPB-MSP domain which carries ALS-causing mutations.

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