Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca

Autores
Droghetti, Enrica; Nicoletti, Francesco Paolo; Bonamore, Alessandra; Boechi, Leonardo; Arroyo Mañez, Pau; Estrin, Dario Ariel; Boffi, Alberto; Smulevich, Giulietta; Feis, Alessandro
Año de publicación
2010
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
An acidic surface variant (ASV) of the "truncated" hemoglobin from Thermobifida fusca was designed with the aim of creating a versatile globin scaffold endowed with thermostability and a high level of recombinant expression in its soluble form while keeping the active site unmodified. This engineered protein was obtained by mutating the surface-exposed residues Phe107 and Arg91 to Glu. Molecular dynamics simulations showed that the mutated residues remain solvent-exposed, not affecting the overall protein structure. Thus, the ASV was used in a combinatorial mutagenesis of the distal heme pocket residues in which one, two, or three of the conserved polar residues [TyrB10(54), TyrCD1(67), and TrpG8(119)] were substituted with Phe. Mutants were characterized by infrared and resonance Raman spectroscopy and compared with the wild-type protein. Similar Fe-proximal His stretching frequencies suggest that none of the mutations alters the proximal side of the heme cavity. Two conformers were observed in the spectra of the CO complexes of both wild-type and ASV protein: form 1 with v(FeC) and v(CO) at 509 and 1938 cm-1 and form 2 with v(FeC) and v(CO) at 518 and 1920 cm-1, respectively. Molecular dynamics simulations were performed for the wild-type and ASV forms, as well as for the TyrB10 mutant. The spectroscopic and computational results demonstrate that CO interacts with TrpG8 in form 1 and interacts with both TrpG8 and TyrCD1 in form 2. TyrB10 does not directly interact with the bound CO.
Fil: Droghetti, Enrica. Universita di Firenze; Italia
Fil: Nicoletti, Francesco Paolo. Universita di Firenze; Italia
Fil: Bonamore, Alessandra. Università degli studi di Roma "La Sapienza"; Italia
Fil: Boechi, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Arroyo Mañez, Pau. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Estrin, Dario Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Boffi, Alberto. Università degli studi di Roma "La Sapienza"; Italia
Fil: Smulevich, Giulietta. Universita di Firenze; Italia
Fil: Feis, Alessandro. Universita di Firenze; Italia
Materia
Structural Properties
Truncated Hemoglobin
Thermofida Fusca
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/83871
Acceder
id CONICETDig_7e9424d78cd9c314d2143feb9fbcd510
oai_identifier_str oai:ri.conicet.gov.ar:11336/83871
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fuscaDroghetti, EnricaNicoletti, Francesco PaoloBonamore, AlessandraBoechi, LeonardoArroyo Mañez, PauEstrin, Dario ArielBoffi, AlbertoSmulevich, GiuliettaFeis, AlessandroStructural PropertiesTruncated HemoglobinThermofida Fuscahttps://purl.org/becyt/ford/1.4https://purl.org/becyt/ford/1An acidic surface variant (ASV) of the "truncated" hemoglobin from Thermobifida fusca was designed with the aim of creating a versatile globin scaffold endowed with thermostability and a high level of recombinant expression in its soluble form while keeping the active site unmodified. This engineered protein was obtained by mutating the surface-exposed residues Phe107 and Arg91 to Glu. Molecular dynamics simulations showed that the mutated residues remain solvent-exposed, not affecting the overall protein structure. Thus, the ASV was used in a combinatorial mutagenesis of the distal heme pocket residues in which one, two, or three of the conserved polar residues [TyrB10(54), TyrCD1(67), and TrpG8(119)] were substituted with Phe. Mutants were characterized by infrared and resonance Raman spectroscopy and compared with the wild-type protein. Similar Fe-proximal His stretching frequencies suggest that none of the mutations alters the proximal side of the heme cavity. Two conformers were observed in the spectra of the CO complexes of both wild-type and ASV protein: form 1 with v(FeC) and v(CO) at 509 and 1938 cm-1 and form 2 with v(FeC) and v(CO) at 518 and 1920 cm-1, respectively. Molecular dynamics simulations were performed for the wild-type and ASV forms, as well as for the TyrB10 mutant. The spectroscopic and computational results demonstrate that CO interacts with TrpG8 in form 1 and interacts with both TrpG8 and TyrCD1 in form 2. TyrB10 does not directly interact with the bound CO.Fil: Droghetti, Enrica. Universita di Firenze; ItaliaFil: Nicoletti, Francesco Paolo. Universita di Firenze; ItaliaFil: Bonamore, Alessandra. Università degli studi di Roma "La Sapienza"; ItaliaFil: Boechi, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; ArgentinaFil: Arroyo Mañez, Pau. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; ArgentinaFil: Estrin, Dario Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; ArgentinaFil: Boffi, Alberto. Università degli studi di Roma "La Sapienza"; ItaliaFil: Smulevich, Giulietta. Universita di Firenze; ItaliaFil: Feis, Alessandro. Universita di Firenze; ItaliaAmerican Chemical Society2010-12info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/83871Droghetti, Enrica; Nicoletti, Francesco Paolo; Bonamore, Alessandra; Boechi, Leonardo; Arroyo Mañez, Pau; et al.; Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca; American Chemical Society; Biochemistry; 49; 49; 12-2010; 10394-104020006-2960CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1021/bi101452kinfo:eu-repo/semantics/altIdentifier/url/https://pubs.acs.org/doi/10.1021/bi101452kinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-10-29T11:42:39Zoai:ri.conicet.gov.ar:11336/83871instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-10-29 11:42:40.142CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
title Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
spellingShingle Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
Droghetti, Enrica
Structural Properties
Truncated Hemoglobin
Thermofida Fusca
title_short Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
title_full Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
title_fullStr Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
title_full_unstemmed Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
title_sort Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca
dc.creator.none.fl_str_mv Droghetti, Enrica
Nicoletti, Francesco Paolo
Bonamore, Alessandra
Boechi, Leonardo
Arroyo Mañez, Pau
Estrin, Dario Ariel
Boffi, Alberto
Smulevich, Giulietta
Feis, Alessandro
author Droghetti, Enrica
author_facet Droghetti, Enrica
Nicoletti, Francesco Paolo
Bonamore, Alessandra
Boechi, Leonardo
Arroyo Mañez, Pau
Estrin, Dario Ariel
Boffi, Alberto
Smulevich, Giulietta
Feis, Alessandro
author_role author
author2 Nicoletti, Francesco Paolo
Bonamore, Alessandra
Boechi, Leonardo
Arroyo Mañez, Pau
Estrin, Dario Ariel
Boffi, Alberto
Smulevich, Giulietta
Feis, Alessandro
author2_role author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv Structural Properties
Truncated Hemoglobin
Thermofida Fusca
topic Structural Properties
Truncated Hemoglobin
Thermofida Fusca
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.4
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv An acidic surface variant (ASV) of the "truncated" hemoglobin from Thermobifida fusca was designed with the aim of creating a versatile globin scaffold endowed with thermostability and a high level of recombinant expression in its soluble form while keeping the active site unmodified. This engineered protein was obtained by mutating the surface-exposed residues Phe107 and Arg91 to Glu. Molecular dynamics simulations showed that the mutated residues remain solvent-exposed, not affecting the overall protein structure. Thus, the ASV was used in a combinatorial mutagenesis of the distal heme pocket residues in which one, two, or three of the conserved polar residues [TyrB10(54), TyrCD1(67), and TrpG8(119)] were substituted with Phe. Mutants were characterized by infrared and resonance Raman spectroscopy and compared with the wild-type protein. Similar Fe-proximal His stretching frequencies suggest that none of the mutations alters the proximal side of the heme cavity. Two conformers were observed in the spectra of the CO complexes of both wild-type and ASV protein: form 1 with v(FeC) and v(CO) at 509 and 1938 cm-1 and form 2 with v(FeC) and v(CO) at 518 and 1920 cm-1, respectively. Molecular dynamics simulations were performed for the wild-type and ASV forms, as well as for the TyrB10 mutant. The spectroscopic and computational results demonstrate that CO interacts with TrpG8 in form 1 and interacts with both TrpG8 and TyrCD1 in form 2. TyrB10 does not directly interact with the bound CO.
Fil: Droghetti, Enrica. Universita di Firenze; Italia
Fil: Nicoletti, Francesco Paolo. Universita di Firenze; Italia
Fil: Bonamore, Alessandra. Università degli studi di Roma "La Sapienza"; Italia
Fil: Boechi, Leonardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Arroyo Mañez, Pau. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Estrin, Dario Ariel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química, Física de los Materiales, Medioambiente y Energía. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química, Física de los Materiales, Medioambiente y Energía; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Inorgánica, Analítica y Química Física; Argentina
Fil: Boffi, Alberto. Università degli studi di Roma "La Sapienza"; Italia
Fil: Smulevich, Giulietta. Universita di Firenze; Italia
Fil: Feis, Alessandro. Universita di Firenze; Italia
description An acidic surface variant (ASV) of the "truncated" hemoglobin from Thermobifida fusca was designed with the aim of creating a versatile globin scaffold endowed with thermostability and a high level of recombinant expression in its soluble form while keeping the active site unmodified. This engineered protein was obtained by mutating the surface-exposed residues Phe107 and Arg91 to Glu. Molecular dynamics simulations showed that the mutated residues remain solvent-exposed, not affecting the overall protein structure. Thus, the ASV was used in a combinatorial mutagenesis of the distal heme pocket residues in which one, two, or three of the conserved polar residues [TyrB10(54), TyrCD1(67), and TrpG8(119)] were substituted with Phe. Mutants were characterized by infrared and resonance Raman spectroscopy and compared with the wild-type protein. Similar Fe-proximal His stretching frequencies suggest that none of the mutations alters the proximal side of the heme cavity. Two conformers were observed in the spectra of the CO complexes of both wild-type and ASV protein: form 1 with v(FeC) and v(CO) at 509 and 1938 cm-1 and form 2 with v(FeC) and v(CO) at 518 and 1920 cm-1, respectively. Molecular dynamics simulations were performed for the wild-type and ASV forms, as well as for the TyrB10 mutant. The spectroscopic and computational results demonstrate that CO interacts with TrpG8 in form 1 and interacts with both TrpG8 and TyrCD1 in form 2. TyrB10 does not directly interact with the bound CO.
publishDate 2010
dc.date.none.fl_str_mv 2010-12
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/83871
Droghetti, Enrica; Nicoletti, Francesco Paolo; Bonamore, Alessandra; Boechi, Leonardo; Arroyo Mañez, Pau; et al.; Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca; American Chemical Society; Biochemistry; 49; 49; 12-2010; 10394-10402
0006-2960
CONICET Digital
CONICET
url http://hdl.handle.net/11336/83871
identifier_str_mv Droghetti, Enrica; Nicoletti, Francesco Paolo; Bonamore, Alessandra; Boechi, Leonardo; Arroyo Mañez, Pau; et al.; Heme pocket structural properties of a bacterial truncated hemoglobin from thermobifida fusca; American Chemical Society; Biochemistry; 49; 49; 12-2010; 10394-10402
0006-2960
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1021/bi101452k
info:eu-repo/semantics/altIdentifier/url/https://pubs.acs.org/doi/10.1021/bi101452k
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv American Chemical Society
publisher.none.fl_str_mv American Chemical Society
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.10058

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