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Title: Interaction of Product Analogues With the Active Site of Rhodobacter Sphaeroides Dimethyl Sulfoxide Reductase

Abstract

We report a structural characterization using X-ray absorption spectroscopy of Rhodobacter sphaeroides dimethylsulfoxide (DMSO) reductase reduced with trimethylarsine, and show that this is structurally analogous to the physiologically relevant dimethylsulfide-reduced DMSO reductase. Our data unambiguously indicate that these species should be regarded as formal MoIV species, and indicate a classical coordination complex of trimethylarsine oxide, with no special structural distortions. The similarity of the trimethylarsine and dimethylsulfide complexes suggests in turn that the dimethylsulfide reduced enzyme possesses a classical coordination of DMSO with no special elongation of the S-O bond, as previously suggested.

Authors:
; ; ; ; ;
Publication Date:
Research Org.:
Stanford Linear Accelerator Center (SLAC)
Sponsoring Org.:
USDOE
OSTI Identifier:
909800
Report Number(s):
SLAC-REPRINT-2007-094
Journal ID: ISSN 0020-1669; INOCAJ; TRN: US200723%%193
DOE Contract Number:
AC02-76SF00515
Resource Type:
Journal Article
Resource Relation:
Journal Name: Inorg. Chem 46:3097,2007; Journal Volume: 46
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ABSORPTION SPECTROSCOPY; DIMETHYL SULFIDE; DMSO; ELONGATION; ENZYMES; OXIDOREDUCTASES; Other,OTHER

Citation Formats

George, G.N., Nelson, K.J., Harris, H.H., Doonan, C.J., Rajagopalan, K.V., and /Saskatchewan U. /Duke U. /Sydney U.. Interaction of Product Analogues With the Active Site of Rhodobacter Sphaeroides Dimethyl Sulfoxide Reductase. United States: N. p., 2007. Web. doi:10.1021/ic061695t.
George, G.N., Nelson, K.J., Harris, H.H., Doonan, C.J., Rajagopalan, K.V., & /Saskatchewan U. /Duke U. /Sydney U.. Interaction of Product Analogues With the Active Site of Rhodobacter Sphaeroides Dimethyl Sulfoxide Reductase. United States. doi:10.1021/ic061695t.
George, G.N., Nelson, K.J., Harris, H.H., Doonan, C.J., Rajagopalan, K.V., and /Saskatchewan U. /Duke U. /Sydney U.. Mon . "Interaction of Product Analogues With the Active Site of Rhodobacter Sphaeroides Dimethyl Sulfoxide Reductase". United States. doi:10.1021/ic061695t.
@article{osti_909800,
title = {Interaction of Product Analogues With the Active Site of Rhodobacter Sphaeroides Dimethyl Sulfoxide Reductase},
author = {George, G.N. and Nelson, K.J. and Harris, H.H. and Doonan, C.J. and Rajagopalan, K.V. and /Saskatchewan U. /Duke U. /Sydney U.},
abstractNote = {We report a structural characterization using X-ray absorption spectroscopy of Rhodobacter sphaeroides dimethylsulfoxide (DMSO) reductase reduced with trimethylarsine, and show that this is structurally analogous to the physiologically relevant dimethylsulfide-reduced DMSO reductase. Our data unambiguously indicate that these species should be regarded as formal MoIV species, and indicate a classical coordination complex of trimethylarsine oxide, with no special structural distortions. The similarity of the trimethylarsine and dimethylsulfide complexes suggests in turn that the dimethylsulfide reduced enzyme possesses a classical coordination of DMSO with no special elongation of the S-O bond, as previously suggested.},
doi = {10.1021/ic061695t},
journal = {Inorg. Chem 46:3097,2007},
number = ,
volume = 46,
place = {United States},
year = {Mon Jul 09 00:00:00 EDT 2007},
month = {Mon Jul 09 00:00:00 EDT 2007}
}
  • X-ray absorption spectroscopy at the molybdenum K-edge has been used to probe the molybdenum coordination of Rhodobacter sphaeroides dimethyl sulfoxide reductase. The molybdenum site of the oxidized protein possesses a novel Mo(VI) mono-oxo site (Mo=O at 1.68 A) with additional coordination by approximately four thiolate ligands at 2.44 A and probably one oxygen or nitogen at 1.92 A. The reduced Mo(IV) form of the enzyme is a des-oxomolybdenum with 3-4 thiolates at 2.33 A and two different Mo-O/N ligands at 2.16 A and 1.92 A. Similarly, the stable Mo(V) glycerol-inhibited species is found to be des-oxomolybdenum with approximately four thiolatemore » ligands at 2.40 A and (probably) two similarly coordinated oxygen or nitrogen ligands at 1.96 A. 43 refs., 5 figs., 1 tab.« less
  • Dimethyl sulfoxide is the substrate to the molybdenum-dependent enzyme dimethyl sulfoxide reductase, which is a member of the large group of molybdenum-containing non-nitrogenase redox enzymes. The active site of these enzymes is thought to possess a so-called molybdopterin, a hydrogenated pterin with an unusual side chain containing a dithiolene group. Up to now the enzyme reactivity was mostly attributed to molybdenum and to the coordination of these sulfur ligands in the side chain. The pterin moiety was not taken into account as playing an active part essential for the enzyme reaction. We demonstrated for the first time a possible coordinationmore » of a hydrogenated pterin to molybdenum with a complex of quinonoid-dihydro-L-biopterin bound to molybdenum in the oxidation state + IV. Now we report the synthesis, crystal structure, and spectroscopic data for trichloro-(quinonoid-N(8)H-6,7-dihydropterin)oxomolybdenum(IV), [MoOCl{sub 3}(H{sup +}-q-H{sub 2}Ptr)](1) (dihydropterin = H{sub 2}Ptr). Crystal data: a = 9.966(3) {angstrom}, b = 14.408(4) {angstrom}, c = 17.362(5) {angstrom}, V = 2493(2) {angstrom}{sup 3}, Z = 8, orthohombic, space group Pbca, R{sub 1} = 0.059 and wR{sub 2} = 0.0150. 1 is synthesized in a redox reaction between Mo(VI)O{sub 2}Cl{sub 2} and tetrahydropterin [H{sub 4}Ptr{center_dot}2HCl] and contains a cationic quinonoid dihydropterin coordinated via the N(5) and O(4) atoms to the molybdenum atom. The crystal structure of 1 containing the hydrogenated pterin exhibits an unusually short Mo-N(5) bond length of 2.013(3) {angstrom}, as compared to 2.324(6) {angstrom} for the corresponding bond in oxidized pterin. 1 is able to quantitatively reduce the substrate dimethyl sulfoxide to dimethyl sulfide under the strict exclusion of oxygen. This reaction can be monitored by {sup 13}C-NMR spectroscopy. A simplified in vivo reaction cycle for the enzyme center of DMSO reductase is proposed as a working hypothesis.« less
  • Structural studies of dimethyl sulfoxide (DMSO) reductases were hampered by modification of the active site during purification. We report an X-ray absorption spectroscopic analysis of the molybdenum active site of Escherichia coli DMSO reductase contained within its native membranes. The enzyme in these preparations is expected to be very close to the form found in vivo. The oxidized active site was found to have four Mo-S ligands at 2.43 angstroms, one Mo=O at 1.71 angstroms, and a longer Mo-O at 1.90 angstroms. We conclude that the oxidized enzyme is a monooxomolybdenum(VI) species coordinated by two molybdopterin dithiolenes and a serine.more » The bond lengths determined for E. coli DMSO reductase are very similar to those determined for the well-characterized Rhodobacter sphaeroides DMSO reductase, suggesting similar active site structures for the two enzymes. Furthermore, our results suggest that the form found in vivo is the monooxobis(molybdopterin) species.« less
  • Molybdenum K-edge X-ray absorption and Mo(V) electron paramagnetic resonance (EPR) spectroscopies have been used to probe the metal coordination in oxidized and reduced forms of both wild-type and a site-directed mutant of Rhodobacter sphaeroides dimethyl sulfoxide (DMSO) reductase. This study confirms earlier findings (George, G.N.; Hilton, J.; Rajagopalan, K.V. J. Am. Chem. Soc. 118, 1113--1117) that the molybdenum site of the oxidized Mo(VI) enzyme possesses one terminal oxygen ligand (Mo{double{underscore}bond}O) at 1.68 {angstrom}, four thiolate ligands at 2.44 {angstrom}, and one oxygen at 1.92{angstrom} and that the dithionite-reduced Mo(IV) enzyme possesses a desoxo species with three or four Mo-S atmore » 2.33 {angstrom} and two different Mo-O ligands at 2.16 and 1.92{angstrom}. Mo(V) EPR indicates the presence of one exchangeable oxygen ligand, most likely an Mo-OH, in the signal-giving species, probably originating from the Mo{double{underscore}bond}O of the oxidized enzyme (E{sub m8.5}(IV/V) = +37 mV, E{sub m8.5}(V/VI) = + 83 mV). The addition of dimethyl sulfide, in the reverse of the physiological reaction, reduces the enzyme. In this form, the enzyme contains a desoxo active site with four Mo-S at 2.36 {angstrom} and two different Mo-O ligands at 1.94 and 2.14{angstrom}. Recombination wild-type R. sphaeroides DMSO reductase expressed in Escherichia coli initially has a dioxo structure (two Mo{double{underscore}bond}O at 1.72 {angstrom} and four Mo-S at 2.48 {angstrom}) but assumes the wild-type Mo(VI) structure after a cycle of reduction and reoxidation. The site-directed Ser147 {r{underscore}arrow} Cys mutant possesses a mono oxo active site in the oxidized state (Mo{double{underscore}bond}O at 1.70 {angstrom}) with five sulfur ligands (at 2.40 {angstrom}), consistent with cysteine 147 coordination to Mo. the dithionite reduced form of the mutant possesses a desoxo site also with five Mo-S ligands (at 2.37 {angstrom}) and one Mo-O at 2.12 {angstrom}. The variant has substantially different Mo(V) EPR and electrochemistry (E{sub m8.5}(IV/V) = {minus}43 mV, E{sub m8.5}(V/VI) = + 106 mV). The active-site structure and catalytic mechanism of DMSO reductase are discussed in the light of these results.« less