Controlling ligand binding in myoglobin by mutagenesis.
A quadruple mutant of sperm whale myoglobin was constructed to mimic the structure found in Ascaris suum hemoglobin. The replacements include His(E7)--Gln, Leu(B10)--Tyr, Thr(E10)--Arg, and Ile(G8)--Phe. Single, double, and triple mutants were characterized to dissect out the effects of the individu...
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2002
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Online Access: | https://www.openaccessrepository.it/record/94547 https://doi.org/10.1074/jbc.m109206200 |
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ftopenaccessrep:oai:zenodo.org:94547 2023-10-25T01:44:08+02:00 Controlling ligand binding in myoglobin by mutagenesis. John S. Olson Federica Draghi Beatrice Vallone Adriana E. Miele Carlo Travaglini-Allocatelli Quentin H. Gibson Maurizio Brunori 2002-03-01 https://www.openaccessrepository.it/record/94547 https://doi.org/10.1074/jbc.m109206200 eng eng url:https://www.openaccessrepository.it/communities/itmirror https://www.openaccessrepository.it/record/94547 doi:10.1074/jbc.m109206200 info:eu-repo/semantics/openAccess https://creativecommons.org/licenses/by/4.0/ Cell Biology Molecular Biology Biochemistry info:eu-repo/semantics/article publication-article 2002 ftopenaccessrep https://doi.org/10.1074/jbc.m109206200 2023-09-26T22:18:22Z A quadruple mutant of sperm whale myoglobin was constructed to mimic the structure found in Ascaris suum hemoglobin. The replacements include His(E7)--Gln, Leu(B10)--Tyr, Thr(E10)--Arg, and Ile(G8)--Phe. Single, double, and triple mutants were characterized to dissect out the effects of the individual substitutions. The crystal structures of the deoxy and oxy forms of the quadruple mutant were determined and compared with that of native Ascaris hemoglobin. Tyr(B10) myoglobin displays low O(2) affinity, high dissociation rate constants, and heterogeneous kinetic behavior, suggesting unfavorable steric interactions between the B10 phenol side chain and His(E7). In contrast, all mutants containing the Tyr(B10)/Gln(E7) pair show high O(2) affinity, low dissociation rate constants, and simple, monophasic kinetic behavior. Replacement of Ile(107) with Phe enhances nanosecond geminate recombination singly and in combination with the Tyr(B10)/Gln(E7)/Arg(E10) mutation by limiting access to the Xe4 site. These kinetic results and comparisons with native Ascaris hemoglobin demonstrate the importance of distal pocket cavities in governing the kinetics of ligand binding. The approximately 150-fold higher O(2) affinity of Ascaris hemoglobin compared with that for Tyr(B10)/Gln(E7)-containing myoglobin mutants appears to be the result of favorable proximal effects in the Ascaris protein, due to a staggered orientation of His(F8), the lack of a hydrogen bonding lattice between the F4, F7, and F8 residues, and the presence of a large polar Trp(G5) residue in the interior portion of the proximal heme pocket. Article in Journal/Newspaper Sperm whale Istituto Nazionale di Fisica Nucleare (INFN): Open Access Repository Journal of Biological Chemistry 277 9 7509 7519 |
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Open Polar |
collection |
Istituto Nazionale di Fisica Nucleare (INFN): Open Access Repository |
op_collection_id |
ftopenaccessrep |
language |
English |
topic |
Cell Biology Molecular Biology Biochemistry |
spellingShingle |
Cell Biology Molecular Biology Biochemistry John S. Olson Federica Draghi Beatrice Vallone Adriana E. Miele Carlo Travaglini-Allocatelli Quentin H. Gibson Maurizio Brunori Controlling ligand binding in myoglobin by mutagenesis. |
topic_facet |
Cell Biology Molecular Biology Biochemistry |
description |
A quadruple mutant of sperm whale myoglobin was constructed to mimic the structure found in Ascaris suum hemoglobin. The replacements include His(E7)--Gln, Leu(B10)--Tyr, Thr(E10)--Arg, and Ile(G8)--Phe. Single, double, and triple mutants were characterized to dissect out the effects of the individual substitutions. The crystal structures of the deoxy and oxy forms of the quadruple mutant were determined and compared with that of native Ascaris hemoglobin. Tyr(B10) myoglobin displays low O(2) affinity, high dissociation rate constants, and heterogeneous kinetic behavior, suggesting unfavorable steric interactions between the B10 phenol side chain and His(E7). In contrast, all mutants containing the Tyr(B10)/Gln(E7) pair show high O(2) affinity, low dissociation rate constants, and simple, monophasic kinetic behavior. Replacement of Ile(107) with Phe enhances nanosecond geminate recombination singly and in combination with the Tyr(B10)/Gln(E7)/Arg(E10) mutation by limiting access to the Xe4 site. These kinetic results and comparisons with native Ascaris hemoglobin demonstrate the importance of distal pocket cavities in governing the kinetics of ligand binding. The approximately 150-fold higher O(2) affinity of Ascaris hemoglobin compared with that for Tyr(B10)/Gln(E7)-containing myoglobin mutants appears to be the result of favorable proximal effects in the Ascaris protein, due to a staggered orientation of His(F8), the lack of a hydrogen bonding lattice between the F4, F7, and F8 residues, and the presence of a large polar Trp(G5) residue in the interior portion of the proximal heme pocket. |
format |
Article in Journal/Newspaper |
author |
John S. Olson Federica Draghi Beatrice Vallone Adriana E. Miele Carlo Travaglini-Allocatelli Quentin H. Gibson Maurizio Brunori |
author_facet |
John S. Olson Federica Draghi Beatrice Vallone Adriana E. Miele Carlo Travaglini-Allocatelli Quentin H. Gibson Maurizio Brunori |
author_sort |
John S. Olson |
title |
Controlling ligand binding in myoglobin by mutagenesis. |
title_short |
Controlling ligand binding in myoglobin by mutagenesis. |
title_full |
Controlling ligand binding in myoglobin by mutagenesis. |
title_fullStr |
Controlling ligand binding in myoglobin by mutagenesis. |
title_full_unstemmed |
Controlling ligand binding in myoglobin by mutagenesis. |
title_sort |
controlling ligand binding in myoglobin by mutagenesis. |
publishDate |
2002 |
url |
https://www.openaccessrepository.it/record/94547 https://doi.org/10.1074/jbc.m109206200 |
genre |
Sperm whale |
genre_facet |
Sperm whale |
op_relation |
url:https://www.openaccessrepository.it/communities/itmirror https://www.openaccessrepository.it/record/94547 doi:10.1074/jbc.m109206200 |
op_rights |
info:eu-repo/semantics/openAccess https://creativecommons.org/licenses/by/4.0/ |
op_doi |
https://doi.org/10.1074/jbc.m109206200 |
container_title |
Journal of Biological Chemistry |
container_volume |
277 |
container_issue |
9 |
container_start_page |
7509 |
op_container_end_page |
7519 |
_version_ |
1780741544318337024 |