Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine
Protein engineering and cofactor replacement have been employed as tools to introduce/modulate peroxidase activity in sperm whale Mb (myoglobin). Based on the rationale that haem peroxidase active sites are characterized by specific charged residues, the Mb haem crevice has been modified to host a h...
Published in: | Biochemical Journal |
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Online Access: | http://hdl.handle.net/11381/1443200 https://doi.org/10.1042/BJ20030863 |
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ftunivparmairis:oai:air.unipr.it:11381/1443200 2024-04-21T08:12:22+00:00 Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine R. RONCONE E. MONZANI M. MURTAS G. BATTAINI A. PENNATI SANANGELANTONI, Anna Maria S. ZUCCOTTI M. BOLOGNESI L. CASELLA R., Roncone E., Monzani M., Murta G., Battaini A., Pennati Sanangelantoni, Anna Maria S., Zuccotti M., Bolognesi L., Casella 2004 http://hdl.handle.net/11381/1443200 https://doi.org/10.1042/BJ20030863 eng eng info:eu-repo/semantics/altIdentifier/wos/WOS:000188927900019 volume:377 firstpage:717 lastpage:724 numberofpages:8 journal:BIOCHEMICAL JOURNAL http://hdl.handle.net/11381/1443200 doi:10.1042/BJ20030863 info:eu-repo/semantics/altIdentifier/scopus/2-s2.0-1242292278 info:eu-repo/semantics/closedAccess info:eu-repo/semantics/article 2004 ftunivparmairis https://doi.org/10.1042/BJ20030863 2024-03-28T01:22:52Z Protein engineering and cofactor replacement have been employed as tools to introduce/modulate peroxidase activity in sperm whale Mb (myoglobin). Based on the rationale that haem peroxidase active sites are characterized by specific charged residues, the Mb haem crevice has been modified to host a haemdistal Arg residue and a proximal Asp, yielding the T67R/S92D Mb mutant. To code extra conformational mobility around the haem, and to increase the peroxidase catalytic efficiency, the T67R/S92D Mb mutant has been subsequently reconstituted with protohaem-L-histidine methyl ester, yielding a stable derivative, T67R/S92D Mb-H. The crystal structure of T67R/S92D cyanometMb (1.4 Å resolution; R factor, 0.12) highlights a regular haem-cyanide binding mode, and the role for themutated residues in affecting the haem propionates as well as the neighbouring water structure. The conformational disorder of the haem propionate- 7 is evidenced by the NMR spectrum of the mutant. Ligandbinding studies show that the iron(III) centres of T67R/S92D Mb, and especially of T67R/S92D Mb-H, exhibit higher affinity for azide and imidazole than wild-type Mb. In addition, both protein derivatives react faster than wild-type Mb with hydrogen peroxide, showing higher peroxidase-like activity towards phenolic substrates. The catalytic efficiency of T67R/S92D Mb-H in these reactions is the highest so far reported for Mb derivatives. A model for the protein–substrate interaction is deduced based on the crystal structure and on the NMR spectra of protein–phenol complexes. Key words: hydrogen peroxide activation, myoglobin mutant,NMR spectra, peroxidase activity, X-ray structure. Article in Journal/Newspaper Sperm whale Archivio della ricerca dell'Università di Parma (CINECA IRIS) Biochemical Journal 377 3 717 724 |
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Open Polar |
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Archivio della ricerca dell'Università di Parma (CINECA IRIS) |
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ftunivparmairis |
language |
English |
description |
Protein engineering and cofactor replacement have been employed as tools to introduce/modulate peroxidase activity in sperm whale Mb (myoglobin). Based on the rationale that haem peroxidase active sites are characterized by specific charged residues, the Mb haem crevice has been modified to host a haemdistal Arg residue and a proximal Asp, yielding the T67R/S92D Mb mutant. To code extra conformational mobility around the haem, and to increase the peroxidase catalytic efficiency, the T67R/S92D Mb mutant has been subsequently reconstituted with protohaem-L-histidine methyl ester, yielding a stable derivative, T67R/S92D Mb-H. The crystal structure of T67R/S92D cyanometMb (1.4 Å resolution; R factor, 0.12) highlights a regular haem-cyanide binding mode, and the role for themutated residues in affecting the haem propionates as well as the neighbouring water structure. The conformational disorder of the haem propionate- 7 is evidenced by the NMR spectrum of the mutant. Ligandbinding studies show that the iron(III) centres of T67R/S92D Mb, and especially of T67R/S92D Mb-H, exhibit higher affinity for azide and imidazole than wild-type Mb. In addition, both protein derivatives react faster than wild-type Mb with hydrogen peroxide, showing higher peroxidase-like activity towards phenolic substrates. The catalytic efficiency of T67R/S92D Mb-H in these reactions is the highest so far reported for Mb derivatives. A model for the protein–substrate interaction is deduced based on the crystal structure and on the NMR spectra of protein–phenol complexes. Key words: hydrogen peroxide activation, myoglobin mutant,NMR spectra, peroxidase activity, X-ray structure. |
author2 |
R., Roncone E., Monzani M., Murta G., Battaini A., Pennati Sanangelantoni, Anna Maria S., Zuccotti M., Bolognesi L., Casella |
format |
Article in Journal/Newspaper |
author |
R. RONCONE E. MONZANI M. MURTAS G. BATTAINI A. PENNATI SANANGELANTONI, Anna Maria S. ZUCCOTTI M. BOLOGNESI L. CASELLA |
spellingShingle |
R. RONCONE E. MONZANI M. MURTAS G. BATTAINI A. PENNATI SANANGELANTONI, Anna Maria S. ZUCCOTTI M. BOLOGNESI L. CASELLA Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
author_facet |
R. RONCONE E. MONZANI M. MURTAS G. BATTAINI A. PENNATI SANANGELANTONI, Anna Maria S. ZUCCOTTI M. BOLOGNESI L. CASELLA |
author_sort |
R. RONCONE |
title |
Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
title_short |
Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
title_full |
Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
title_fullStr |
Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
title_full_unstemmed |
Engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the T67R/S92D mutant and its derivative reconstituted with protohemin-L-histidine |
title_sort |
engineering peroxidase activity in myoglobin: the heme cavity structure and peroxide activation in the t67r/s92d mutant and its derivative reconstituted with protohemin-l-histidine |
publishDate |
2004 |
url |
http://hdl.handle.net/11381/1443200 https://doi.org/10.1042/BJ20030863 |
genre |
Sperm whale |
genre_facet |
Sperm whale |
op_relation |
info:eu-repo/semantics/altIdentifier/wos/WOS:000188927900019 volume:377 firstpage:717 lastpage:724 numberofpages:8 journal:BIOCHEMICAL JOURNAL http://hdl.handle.net/11381/1443200 doi:10.1042/BJ20030863 info:eu-repo/semantics/altIdentifier/scopus/2-s2.0-1242292278 |
op_rights |
info:eu-repo/semantics/closedAccess |
op_doi |
https://doi.org/10.1042/BJ20030863 |
container_title |
Biochemical Journal |
container_volume |
377 |
container_issue |
3 |
container_start_page |
717 |
op_container_end_page |
724 |
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1796932395431100416 |