Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1
In all structurally characterized bacterial multicomponent monooxygenase (BMM) hydroxylase proteins, a series of hydrophobic cavities in the α-subunit trace a conserved path from the protein exterior to the carboxylate-bridged diiron active site. This study examines these cavities as a potential rou...
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| Format: | Article |
| Language: | en_US |
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American Chemical Society
2013
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| Online Access: | http://hdl.handle.net/1721.1/76212 https://orcid.org/0000-0002-2693-4982 |
| _version_ | 1811089964455165952 |
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| author | McCormick, Michael S. Lippard, Stephen J. |
| author2 | Massachusetts Institute of Technology. Department of Chemistry |
| author_facet | Massachusetts Institute of Technology. Department of Chemistry McCormick, Michael S. Lippard, Stephen J. |
| author_sort | McCormick, Michael S. |
| collection | MIT |
| description | In all structurally characterized bacterial multicomponent monooxygenase (BMM) hydroxylase proteins, a series of hydrophobic cavities in the α-subunit trace a conserved path from the protein exterior to the carboxylate-bridged diiron active site. This study examines these cavities as a potential route for transport of dioxygen to the active site by crystallographic characterization of a xenon-pressurized sample of the hydroxylase component of phenol hydroxylase from Pseudomonas sp. OX1. Computational analyses of the hydrophobic cavities in the hydroxylase α-subunits of phenol hydroxylase (PHH), soluble methane monooxygenase (MMOH), and toluene/o-xylene monooxygenase (ToMOH) are also presented. The results, together with previous findings from crystallographic studies of xenon-pressurized sMMO hydroxylase, clearly identify the propensity for these cavities to bind hydrophobic gas molecules in the protein interior. This proposed functional role is supported by recent stopped flow kinetic studies of ToMOH variants [Song, W. J., et al. (2011) Proc. Natl. Acad. Sci. U.S.A.108, 14795–14800]. In addition to information about the Xe sites, the structure determination revealed significantly weakened binding of regulatory protein to the hydroxylase in comparison to that in the previously reported structure of PHH, as well as the presence of a newly identified metal-binding site in the α-subunit that adopts a linear coordination environment consistent with Cu(I), and a glycerol molecule bound to Fe1 in a fashion that is unique among hydrocarbon–diiron site adducts reported to date in BMM hydroxylase structures. Finally, a comparative analysis of the α-subunit structures of PHH, MMOH, and ToMOH details proposed routes for the other three BMM substrates, the hydrocarbon, electrons, and protons, comprising cavities, channels, hydrogen-bonding networks, and pores in the structures of their α-subunits. |
| first_indexed | 2024-09-23T14:28:29Z |
| format | Article |
| id | mit-1721.1/76212 |
| institution | Massachusetts Institute of Technology |
| language | en_US |
| last_indexed | 2024-09-23T14:28:29Z |
| publishDate | 2013 |
| publisher | American Chemical Society |
| record_format | dspace |
| spelling | mit-1721.1/762122022-09-29T09:36:07Z Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 McCormick, Michael S. Lippard, Stephen J. Massachusetts Institute of Technology. Department of Chemistry Lippard, Stephen J. Lippard, Stephen J. McCormick, Michael S. In all structurally characterized bacterial multicomponent monooxygenase (BMM) hydroxylase proteins, a series of hydrophobic cavities in the α-subunit trace a conserved path from the protein exterior to the carboxylate-bridged diiron active site. This study examines these cavities as a potential route for transport of dioxygen to the active site by crystallographic characterization of a xenon-pressurized sample of the hydroxylase component of phenol hydroxylase from Pseudomonas sp. OX1. Computational analyses of the hydrophobic cavities in the hydroxylase α-subunits of phenol hydroxylase (PHH), soluble methane monooxygenase (MMOH), and toluene/o-xylene monooxygenase (ToMOH) are also presented. The results, together with previous findings from crystallographic studies of xenon-pressurized sMMO hydroxylase, clearly identify the propensity for these cavities to bind hydrophobic gas molecules in the protein interior. This proposed functional role is supported by recent stopped flow kinetic studies of ToMOH variants [Song, W. J., et al. (2011) Proc. Natl. Acad. Sci. U.S.A.108, 14795–14800]. In addition to information about the Xe sites, the structure determination revealed significantly weakened binding of regulatory protein to the hydroxylase in comparison to that in the previously reported structure of PHH, as well as the presence of a newly identified metal-binding site in the α-subunit that adopts a linear coordination environment consistent with Cu(I), and a glycerol molecule bound to Fe1 in a fashion that is unique among hydrocarbon–diiron site adducts reported to date in BMM hydroxylase structures. Finally, a comparative analysis of the α-subunit structures of PHH, MMOH, and ToMOH details proposed routes for the other three BMM substrates, the hydrocarbon, electrons, and protons, comprising cavities, channels, hydrogen-bonding networks, and pores in the structures of their α-subunits. National Institute of General Medical Sciences (U.S.) (Grant GM032134) 2013-01-08T19:41:49Z 2013-01-08T19:41:49Z 2011-12 Article http://purl.org/eprint/type/JournalArticle 0006-2960 1520-4995 http://hdl.handle.net/1721.1/76212 McCormick, Michael S., and Stephen J. Lippard. “Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp. OX1.” Biochemistry 50.51 (2011): 11058–11069. Web. https://orcid.org/0000-0002-2693-4982 en_US http://dx.doi.org/10.1021/bi201248b Biochemistry Article is made available in accordance with the publisher's policy and may be subject to US copyright law. Please refer to the publisher's site for terms of use. application/pdf American Chemical Society Prof. Lippard via Erja Kajosalo |
| spellingShingle | McCormick, Michael S. Lippard, Stephen J. Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title | Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title_full | Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title_fullStr | Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title_full_unstemmed | Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title_short | Analysis of Substrate Access to Active Sites in Bacterial Multicomponent Monooxygenase Hydroxylases: X-Ray Crystal Structure of Xenon-Pressurized Phenol Hydroxylase from Pseudomonas Sp Ox1 |
| title_sort | analysis of substrate access to active sites in bacterial multicomponent monooxygenase hydroxylases x ray crystal structure of xenon pressurized phenol hydroxylase from pseudomonas sp ox1 |
| url | http://hdl.handle.net/1721.1/76212 https://orcid.org/0000-0002-2693-4982 |
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