Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).

Type I restriction-modification (RM) systems are comprised of two multi-subunit enzymes, the methyltransferase (∼160 kDa), responsible for methylation of DNA, and the restriction endonuclease (∼400 kDa), responsible for DNA cleavage. Both enzymes share a number of subunits. An engineered RM system,...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	James E Taylor, Anna Swiderska, Jean-Baptiste Artero, Philip Callow, Geoff Kneale
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/4527021a2dd3489b9d5746d874fa7815
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:4527021a2dd3489b9d5746d874fa7815
record_format	dspace
spelling	oai:doaj.org-article:4527021a2dd3489b9d5746d874fa78152021-11-18T07:22:57ZStructural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).1932-620310.1371/journal.pone.0035263https://doaj.org/article/4527021a2dd3489b9d5746d874fa78152012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22493743/?tool=EBIhttps://doaj.org/toc/1932-6203Type I restriction-modification (RM) systems are comprised of two multi-subunit enzymes, the methyltransferase (∼160 kDa), responsible for methylation of DNA, and the restriction endonuclease (∼400 kDa), responsible for DNA cleavage. Both enzymes share a number of subunits. An engineered RM system, EcoR124I(NT), based on the N-terminal domain of the specificity subunit of EcoR124I was constructed that recognises the symmetrical sequence GAAN(7)TTC and is active as a methyltransferase. Here, we investigate the restriction endonuclease activity of R. EcoR124I(NT)in vitro and the subunit assembly of the multi-subunit enzyme. Finally, using small-angle neutron scattering and selective deuteration, we present a low-resolution structural model of the endonuclease and locate the motor subunits within the multi-subunit enzyme. We show that the covalent linkage between the two target recognition domains of the specificity subunit is not required for subunit assembly or enzyme activity, and discuss the implications for the evolution of Type I enzymes.James E TaylorAnna SwiderskaJean-Baptiste ArteroPhilip CallowGeoff KnealePublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 4, p e35263 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q James E Taylor Anna Swiderska Jean-Baptiste Artero Philip Callow Geoff Kneale Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
description	Type I restriction-modification (RM) systems are comprised of two multi-subunit enzymes, the methyltransferase (∼160 kDa), responsible for methylation of DNA, and the restriction endonuclease (∼400 kDa), responsible for DNA cleavage. Both enzymes share a number of subunits. An engineered RM system, EcoR124I(NT), based on the N-terminal domain of the specificity subunit of EcoR124I was constructed that recognises the symmetrical sequence GAAN(7)TTC and is active as a methyltransferase. Here, we investigate the restriction endonuclease activity of R. EcoR124I(NT)in vitro and the subunit assembly of the multi-subunit enzyme. Finally, using small-angle neutron scattering and selective deuteration, we present a low-resolution structural model of the endonuclease and locate the motor subunits within the multi-subunit enzyme. We show that the covalent linkage between the two target recognition domains of the specificity subunit is not required for subunit assembly or enzyme activity, and discuss the implications for the evolution of Type I enzymes.
format	article
author	James E Taylor Anna Swiderska Jean-Baptiste Artero Philip Callow Geoff Kneale
author_facet	James E Taylor Anna Swiderska Jean-Baptiste Artero Philip Callow Geoff Kneale
author_sort	James E Taylor
title	Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
title_short	Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
title_full	Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
title_fullStr	Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
title_full_unstemmed	Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).
title_sort	structural and functional analysis of the symmetrical type i restriction endonuclease r.ecor124i(nt).
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/4527021a2dd3489b9d5746d874fa7815
work_keys_str_mv	AT jamesetaylor structuralandfunctionalanalysisofthesymmetricaltypeirestrictionendonucleaserecor124int AT annaswiderska structuralandfunctionalanalysisofthesymmetricaltypeirestrictionendonucleaserecor124int AT jeanbaptisteartero structuralandfunctionalanalysisofthesymmetricaltypeirestrictionendonucleaserecor124int AT philipcallow structuralandfunctionalanalysisofthesymmetricaltypeirestrictionendonucleaserecor124int AT geoffkneale structuralandfunctionalanalysisofthesymmetricaltypeirestrictionendonucleaserecor124int
_version_	1718423530619535360

Structural and functional analysis of the symmetrical Type I restriction endonuclease R.EcoR124I(NT).

Ejemplares similares