Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function

10 páginas, 5 figuras, 1 tabla. Contiene figura S1 del material suplementario

Detalles Bibliográficos
Autores: Díez-Fernández, Carmen, Hu, Liyan, Cervera, Javier, Häberle, Johannes, Rubio, Vicente
Tipo de recurso: artículo
Estado:Versión aceptada para publicación
Fecha de publicación:2014
País:España
Institución:Consejo Superior de Investigaciones Científicas (CSIC)
Repositorio:DIGITAL.CSIC. Repositorio Institucional del CSIC
OAI Identifier:oai:digital.csic.es:10261/109003
Acceso en línea:http://hdl.handle.net/10261/109003
Access Level:acceso abierto
Palabra clave:Urea cycle diseases
CPS1 deficiency
Hyperammonemia
Inborn errors
CPS 1 structure
Site-directed mutagenesis
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spelling Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown functionDíez-Fernández, CarmenHu, LiyanCervera, JavierHäberle, JohannesRubio, VicenteUrea cycle diseasesCPS1 deficiencyHyperammonemiaInborn errorsCPS 1 structureSite-directed mutagenesis10 páginas, 5 figuras, 1 tabla. Contiene figura S1 del material suplementarioCarbamoyl phosphate synthetase 1 deficiency (CPS1D) is an inborn error of the urea cycle that is due to mutations in the CPS1 gene. In the first large repertory of mutations found in CPS1D, a small CPS1 domain of unknown function (called the UFSD) was found to host missense changes with high frequency, despite the fact that this domain does not host substrate-binding or catalytic machinery. We investigate here by in vitro expression studies using baculovirus/insect cells the reasons for the prominence of the UFSD in CPS1D, as well as the disease-causing roles and pathogenic mechanisms of the mutations affecting this domain. All but three of the 18 missense changes found thus far mapping in this domain in CPS1D patients drastically decreased the yield of pure CPS1, mainly because of decreased enzyme solubility, strongly suggesting misfolding as a major determinant of the mutations negative effects. In addition, the majority of the mutations also decreased from modestly to very drastically the specific activity of the fraction of the enzyme that remained soluble and that could be purified, apparently because they decreased V(max). Substantial although not dramatic increases in K(m) values for the substrates or for N-acetyl-L-glutamate were observed for only five mutations. Similarly, important thermal stability decreases were observed for three mutations. The results indicate a disease-causing role for all the mutations, due in most cases to the combined effects of the low enzyme level and the decreased activity. Our data strongly support the value of the present expression system for ascertaining the disease-causing potential of CPS1 mutations, provided that the CPS1 yield is monitored. The observed effects of the mutations have been rationalized on the basis of an existing structural model of CPS1. This model shows that the UFSD, which is in the middle of the 1462-residue multidomain CPS1 protein, plays a key integrating role for creating the CPS1 multidomain architecture leading us to propose here a denomination of "Integrating Domain" for this CPS1 region. The majority of these 18 mutations distort the interaction of this domain with other CPS1 domains, in many cases by causing improper folding of structural elements of the Integrating Domain that play key roles in these interactions.This work was supported by grants from the Fundación Alicia Koplowitz, the Valencian and Spanish governments (Prometeo 2009/051 and BFU2011-30407, respectively) and the Swiss National Science Foundation (grant 310030_127184). C.D-F was a FPU fellow of the Spanish Government and received from that Government a bursary for short-time work in Zurich.Peer reviewedElsevierConsejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]201420142014info:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_6501Postprintinfo:eu-repo/semantics/acceptedVersionhttp://hdl.handle.net/10261/109003reponame:DIGITAL.CSIC. Repositorio Institucional del CSICinstname:Consejo Superior de Investigaciones Científicas (CSIC)Ingléshttp://dx.doi.org/10.1016/j.ymgme.2014.04.003Síinfo:eu-repo/semantics/openAccessoai:digital.csic.es:10261/1090032026-05-22T06:33:51Z
dc.title.none.fl_str_mv Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
title Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
spellingShingle Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
Díez-Fernández, Carmen
Urea cycle diseases
CPS1 deficiency
Hyperammonemia
Inborn errors
CPS 1 structure
Site-directed mutagenesis
title_short Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
title_full Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
title_fullStr Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
title_full_unstemmed Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
title_sort Understanding carbamoyl phosphate synthetase (CPS1) deficiency by using the recombinantly purified human enzyme: effects of CPS1 mutations that concentrate in a central domain of unknown function
dc.creator.none.fl_str_mv Díez-Fernández, Carmen
Hu, Liyan
Cervera, Javier
Häberle, Johannes
Rubio, Vicente
author Díez-Fernández, Carmen
author_facet Díez-Fernández, Carmen
Hu, Liyan
Cervera, Javier
Häberle, Johannes
Rubio, Vicente
author_role author
author2 Hu, Liyan
Cervera, Javier
Häberle, Johannes
Rubio, Vicente
author2_role author
author
author
author
dc.contributor.none.fl_str_mv Consejo Superior de Investigaciones Científicas [https://ror.org/02gfc7t72]
dc.subject.none.fl_str_mv Urea cycle diseases
CPS1 deficiency
Hyperammonemia
Inborn errors
CPS 1 structure
Site-directed mutagenesis
topic Urea cycle diseases
CPS1 deficiency
Hyperammonemia
Inborn errors
CPS 1 structure
Site-directed mutagenesis
description 10 páginas, 5 figuras, 1 tabla. Contiene figura S1 del material suplementario
publishDate 2014
dc.date.none.fl_str_mv 2014
2014
2014
dc.type.none.fl_str_mv info:eu-repo/semantics/article
http://purl.org/coar/resource_type/c_6501
Postprint
info:eu-repo/semantics/acceptedVersion
format article
status_str acceptedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/10261/109003
url http://hdl.handle.net/10261/109003
dc.language.none.fl_str_mv Inglés
language_invalid_str_mv Inglés
dc.relation.none.fl_str_mv http://dx.doi.org/10.1016/j.ymgme.2014.04.003
Sí
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
eu_rights_str_mv openAccess
dc.publisher.none.fl_str_mv Elsevier
publisher.none.fl_str_mv Elsevier
dc.source.none.fl_str_mv reponame:DIGITAL.CSIC. Repositorio Institucional del CSIC
instname:Consejo Superior de Investigaciones Científicas (CSIC)
instname_str Consejo Superior de Investigaciones Científicas (CSIC)
reponame_str DIGITAL.CSIC. Repositorio Institucional del CSIC
collection DIGITAL.CSIC. Repositorio Institucional del CSIC
repository.name.fl_str_mv
repository.mail.fl_str_mv
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score 15.812429

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