Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies
The extradiol, aromatic ring-cleaving enzyme homoprotocatechuate 2,3-dioxygenase (HPCD) catalyzes a complex chain of reactions that involve second sphere residues of the active site. The importance of the 2nd-sphere residue His200 was demonstrated in studies of HPCD variants, such as His200Cys (H200...
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ftpubmed:oai:pubmedcentral.nih.gov:4924929 2023-05-15T17:53:58+02:00 Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies Meier, Katlyn K. Rogers, Melanie S. Kovaleva, Elena G. Lipscomb, John D. Bominaar, Emile L. Münck, Eckard 2016-06-08 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4924929/ http://www.ncbi.nlm.nih.gov/pubmed/27275865 https://doi.org/10.1021/acs.inorgchem.6b00148 en eng http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4924929/ http://www.ncbi.nlm.nih.gov/pubmed/27275865 http://dx.doi.org/10.1021/acs.inorgchem.6b00148 Article Text 2016 ftpubmed https://doi.org/10.1021/acs.inorgchem.6b00148 2017-06-24T23:59:42Z The extradiol, aromatic ring-cleaving enzyme homoprotocatechuate 2,3-dioxygenase (HPCD) catalyzes a complex chain of reactions that involve second sphere residues of the active site. The importance of the 2nd-sphere residue His200 was demonstrated in studies of HPCD variants, such as His200Cys (H200C), which revealed significant retardations of certain steps in the catalytic process as a result of the substitution, allowing novel reaction cycle intermediates to be trapped for spectroscopic characterization. As the H200C variant largely retains the wild-type active site structure and produces the correct ring-cleaved product, this variant presents a valuable target for mechanistic HPCD studies. Here the high-spin FeII states of resting H200C and the H200C–homoprotocatechuate enzyme–substrate (ES) complex have been characterized with Mössbauer spectroscopy to assess the electronic structures of the active site in these states. The analysis reveals a high-spin FeII center in a low symmetry environment that is reflected in the values of the zero-field splitting (ZFS) (D ≈ −8 cm−1, E/D ≈ 1/3 in ES) as well as the relative orientations of the principal axes of the 57Fe magnetic hyperfine (A) and electric field gradient (EFG) tensors relative to the ZFS tensor axes. A spin Hamiltonian analysis of the spectra for the ES complex indicates that the magnetization axis of the integer-spin S = 2 FeII system is nearly parallel to the symmetry axis, z, of the doubly occupied dxy ground orbital deduced from the EFG and A-values, an observation which cannot be rationalized by DFT assisted crystal-field theory. In contrast, ORCA/CASSCF calculations for the ZFS tensor in combination with DFT calculations for the EFG- and A-tensors describe the experimental data remarkably well. Text Orca PubMed Central (PMC) Inorganic Chemistry 55 12 5862 5870 |
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Article Meier, Katlyn K. Rogers, Melanie S. Kovaleva, Elena G. Lipscomb, John D. Bominaar, Emile L. Münck, Eckard Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
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The extradiol, aromatic ring-cleaving enzyme homoprotocatechuate 2,3-dioxygenase (HPCD) catalyzes a complex chain of reactions that involve second sphere residues of the active site. The importance of the 2nd-sphere residue His200 was demonstrated in studies of HPCD variants, such as His200Cys (H200C), which revealed significant retardations of certain steps in the catalytic process as a result of the substitution, allowing novel reaction cycle intermediates to be trapped for spectroscopic characterization. As the H200C variant largely retains the wild-type active site structure and produces the correct ring-cleaved product, this variant presents a valuable target for mechanistic HPCD studies. Here the high-spin FeII states of resting H200C and the H200C–homoprotocatechuate enzyme–substrate (ES) complex have been characterized with Mössbauer spectroscopy to assess the electronic structures of the active site in these states. The analysis reveals a high-spin FeII center in a low symmetry environment that is reflected in the values of the zero-field splitting (ZFS) (D ≈ −8 cm−1, E/D ≈ 1/3 in ES) as well as the relative orientations of the principal axes of the 57Fe magnetic hyperfine (A) and electric field gradient (EFG) tensors relative to the ZFS tensor axes. A spin Hamiltonian analysis of the spectra for the ES complex indicates that the magnetization axis of the integer-spin S = 2 FeII system is nearly parallel to the symmetry axis, z, of the doubly occupied dxy ground orbital deduced from the EFG and A-values, an observation which cannot be rationalized by DFT assisted crystal-field theory. In contrast, ORCA/CASSCF calculations for the ZFS tensor in combination with DFT calculations for the EFG- and A-tensors describe the experimental data remarkably well. |
format |
Text |
author |
Meier, Katlyn K. Rogers, Melanie S. Kovaleva, Elena G. Lipscomb, John D. Bominaar, Emile L. Münck, Eckard |
author_facet |
Meier, Katlyn K. Rogers, Melanie S. Kovaleva, Elena G. Lipscomb, John D. Bominaar, Emile L. Münck, Eckard |
author_sort |
Meier, Katlyn K. |
title |
Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
title_short |
Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
title_full |
Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
title_fullStr |
Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
title_full_unstemmed |
Enzyme Substrate Complex of the H200C Variant of Homoprotocatechuate 2,3-Dioxygenase: Mössbauer and Computational Studies |
title_sort |
enzyme substrate complex of the h200c variant of homoprotocatechuate 2,3-dioxygenase: mössbauer and computational studies |
publishDate |
2016 |
url |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4924929/ http://www.ncbi.nlm.nih.gov/pubmed/27275865 https://doi.org/10.1021/acs.inorgchem.6b00148 |
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4924929/ http://www.ncbi.nlm.nih.gov/pubmed/27275865 http://dx.doi.org/10.1021/acs.inorgchem.6b00148 |
op_doi |
https://doi.org/10.1021/acs.inorgchem.6b00148 |
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Inorganic Chemistry |
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55 |
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12 |
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5862 |
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5870 |
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1766161681040277504 |