Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases
- Autores
- Musumeci, Matias Alejandro; Botti, Horacio; Buschiazzo, Alejandro; Ceccarelli, Eduardo Augusto
- Año de publicación
- 2011
- Idioma
- inglés
- Tipo de recurso
- artículo
- Estado
- versión publicada
- Descripción
- Plant-type ferredoxin-NADP(H) reductases (FNRs) are grouped in two classes, plastidic with an extended FAD conformation and high catalytic rates and bacterial with a folded flavin nucleotide and low turnover rates. The 112-123 β-hairpin from a plastidic FNR and the carboxy-terminaltryptophan of a bacterial FNR, suggested to be responsible for the FAD differential conformation, were mutually exchanged. The plastidic FNR lacking the β-hairpin was unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial FNR, resulted in an enzyme withdecreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the β-hairpin into the corresponding position of the bacterialFNR increased FAD affinity but did not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produced a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzyme?s ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras showed no significant changes in their overall structure, although alterations in the FAD conformations were observed. Plastidic and bacterial FNRs thus reveal differential effects of key structural elements. While the 112-123 β-hairpin modulates the catalytic efficiency of plastidic FNR, it seems not to affect the bacterial FNR behavior, which instead can be improved by the loss of the C-terminal tryptophan. This report highlights the role of the FAD moiety conformation and the structural determinants involved in stabilizing it, ultimately modulating the functional output of FNRs.
Fil: Musumeci, Matias Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina
Fil: Botti, Horacio. Instituto Pasteur de Montevideo; Uruguay
Fil: Buschiazzo, Alejandro. Instituto Pasteur de Montevideo; Uruguay
Fil: Ceccarelli, Eduardo Augusto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina - Materia
-
Amino Acid Motifs
Enzyme Stability
Ferredoxin-NADP Reductase
Protein Unfolding - Nivel de accesibilidad
- acceso abierto
- Condiciones de uso
- https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
- Repositorio
- Institución
- Consejo Nacional de Investigaciones Científicas y Técnicas
- OAI Identificador
- oai:ri.conicet.gov.ar:11336/268125
Ver los metadatos del registro completo
id |
CONICETDig_291f51e3e585993fbf26605b9d9aba0c |
---|---|
oai_identifier_str |
oai:ri.conicet.gov.ar:11336/268125 |
network_acronym_str |
CONICETDig |
repository_id_str |
3498 |
network_name_str |
CONICET Digital (CONICET) |
spelling |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) ReductasesMusumeci, Matias AlejandroBotti, HoracioBuschiazzo, AlejandroCeccarelli, Eduardo AugustoAmino Acid MotifsEnzyme StabilityFerredoxin-NADP ReductaseProtein Unfoldinghttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Plant-type ferredoxin-NADP(H) reductases (FNRs) are grouped in two classes, plastidic with an extended FAD conformation and high catalytic rates and bacterial with a folded flavin nucleotide and low turnover rates. The 112-123 β-hairpin from a plastidic FNR and the carboxy-terminaltryptophan of a bacterial FNR, suggested to be responsible for the FAD differential conformation, were mutually exchanged. The plastidic FNR lacking the β-hairpin was unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial FNR, resulted in an enzyme withdecreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the β-hairpin into the corresponding position of the bacterialFNR increased FAD affinity but did not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produced a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzyme?s ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras showed no significant changes in their overall structure, although alterations in the FAD conformations were observed. Plastidic and bacterial FNRs thus reveal differential effects of key structural elements. While the 112-123 β-hairpin modulates the catalytic efficiency of plastidic FNR, it seems not to affect the bacterial FNR behavior, which instead can be improved by the loss of the C-terminal tryptophan. This report highlights the role of the FAD moiety conformation and the structural determinants involved in stabilizing it, ultimately modulating the functional output of FNRs.Fil: Musumeci, Matias Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Botti, Horacio. Instituto Pasteur de Montevideo; UruguayFil: Buschiazzo, Alejandro. Instituto Pasteur de Montevideo; UruguayFil: Ceccarelli, Eduardo Augusto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaAmerican Chemical Society2011-03info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/268125Musumeci, Matias Alejandro; Botti, Horacio; Buschiazzo, Alejandro; Ceccarelli, Eduardo Augusto; Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases; American Chemical Society; Biochemistry; 50; 12; 3-2011; 2111-21220006-2960CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://pubs.acs.org/doi/abs/10.1021/bi101772ainfo:eu-repo/semantics/altIdentifier/doi/10.1021/bi101772ainfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T09:46:53Zoai:ri.conicet.gov.ar:11336/268125instacron: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:34982025-09-29 09:46:53.797CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse |
dc.title.none.fl_str_mv |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
title |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
spellingShingle |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases Musumeci, Matias Alejandro Amino Acid Motifs Enzyme Stability Ferredoxin-NADP Reductase Protein Unfolding |
title_short |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
title_full |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
title_fullStr |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
title_full_unstemmed |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
title_sort |
Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases |
dc.creator.none.fl_str_mv |
Musumeci, Matias Alejandro Botti, Horacio Buschiazzo, Alejandro Ceccarelli, Eduardo Augusto |
author |
Musumeci, Matias Alejandro |
author_facet |
Musumeci, Matias Alejandro Botti, Horacio Buschiazzo, Alejandro Ceccarelli, Eduardo Augusto |
author_role |
author |
author2 |
Botti, Horacio Buschiazzo, Alejandro Ceccarelli, Eduardo Augusto |
author2_role |
author author author |
dc.subject.none.fl_str_mv |
Amino Acid Motifs Enzyme Stability Ferredoxin-NADP Reductase Protein Unfolding |
topic |
Amino Acid Motifs Enzyme Stability Ferredoxin-NADP Reductase Protein Unfolding |
purl_subject.fl_str_mv |
https://purl.org/becyt/ford/1.6 https://purl.org/becyt/ford/1 |
dc.description.none.fl_txt_mv |
Plant-type ferredoxin-NADP(H) reductases (FNRs) are grouped in two classes, plastidic with an extended FAD conformation and high catalytic rates and bacterial with a folded flavin nucleotide and low turnover rates. The 112-123 β-hairpin from a plastidic FNR and the carboxy-terminaltryptophan of a bacterial FNR, suggested to be responsible for the FAD differential conformation, were mutually exchanged. The plastidic FNR lacking the β-hairpin was unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial FNR, resulted in an enzyme withdecreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the β-hairpin into the corresponding position of the bacterialFNR increased FAD affinity but did not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produced a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzyme?s ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras showed no significant changes in their overall structure, although alterations in the FAD conformations were observed. Plastidic and bacterial FNRs thus reveal differential effects of key structural elements. While the 112-123 β-hairpin modulates the catalytic efficiency of plastidic FNR, it seems not to affect the bacterial FNR behavior, which instead can be improved by the loss of the C-terminal tryptophan. This report highlights the role of the FAD moiety conformation and the structural determinants involved in stabilizing it, ultimately modulating the functional output of FNRs. Fil: Musumeci, Matias Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina Fil: Botti, Horacio. Instituto Pasteur de Montevideo; Uruguay Fil: Buschiazzo, Alejandro. Instituto Pasteur de Montevideo; Uruguay Fil: Ceccarelli, Eduardo Augusto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; Argentina |
description |
Plant-type ferredoxin-NADP(H) reductases (FNRs) are grouped in two classes, plastidic with an extended FAD conformation and high catalytic rates and bacterial with a folded flavin nucleotide and low turnover rates. The 112-123 β-hairpin from a plastidic FNR and the carboxy-terminaltryptophan of a bacterial FNR, suggested to be responsible for the FAD differential conformation, were mutually exchanged. The plastidic FNR lacking the β-hairpin was unable to fold properly. An extra tryptophan at the carboxy terminus, emulating the bacterial FNR, resulted in an enzyme withdecreased affinity for FAD and reduced diaphorase and ferredoxin-dependent cytochrome c reductase activities. The insertion of the β-hairpin into the corresponding position of the bacterialFNR increased FAD affinity but did not affect its catalytic properties. The same insertion with simultaneous deletion of the carboxyterminal tryptophan produced a bacterial chimera emulating the plastidic architecture with an increased kcat and an increased catalytic efficiency for the diaphorase activity and a decrease in the enzyme?s ability to react with its substrates ferredoxin and flavodoxin. Crystallographic structures of the chimeras showed no significant changes in their overall structure, although alterations in the FAD conformations were observed. Plastidic and bacterial FNRs thus reveal differential effects of key structural elements. While the 112-123 β-hairpin modulates the catalytic efficiency of plastidic FNR, it seems not to affect the bacterial FNR behavior, which instead can be improved by the loss of the C-terminal tryptophan. This report highlights the role of the FAD moiety conformation and the structural determinants involved in stabilizing it, ultimately modulating the functional output of FNRs. |
publishDate |
2011 |
dc.date.none.fl_str_mv |
2011-03 |
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/11336/268125 Musumeci, Matias Alejandro; Botti, Horacio; Buschiazzo, Alejandro; Ceccarelli, Eduardo Augusto; Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases; American Chemical Society; Biochemistry; 50; 12; 3-2011; 2111-2122 0006-2960 CONICET Digital CONICET |
url |
http://hdl.handle.net/11336/268125 |
identifier_str_mv |
Musumeci, Matias Alejandro; Botti, Horacio; Buschiazzo, Alejandro; Ceccarelli, Eduardo Augusto; Swapping FAD Binding Motifs between Plastidic and Bacterial Ferredoxin-NADP(H) Reductases; American Chemical Society; Biochemistry; 50; 12; 3-2011; 2111-2122 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/url/http://pubs.acs.org/doi/abs/10.1021/bi101772a info:eu-repo/semantics/altIdentifier/doi/10.1021/bi101772a |
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 |
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 |
_version_ |
1844613462816194560 |
score |
13.070432 |