Oxo-carboxylato-molybdenum(VI) complexes possessing dithiolene ligands related to the active site of type II DMSOR family molybdoenzymes

Abstract

Spectroscopic and kinetic studies indicate that oxo-carboxylato-molybdenum(VI) bis-dithiolene complexes, (Mo^VIO(p-X-OBz)L₂), have been generated at low temperature as active site structural models for the type II class of pyranopterin molybdenum DMSOR family enzymes. A DFT analysis of low energy charge transfer bands shows that these complexes possess a Mo–S_dithiolene π-bonding interaction between the Mo(d_xy) redox active molecular orbital and a cis S(p_z) donor orbital located on one of the dithiolene ligands.

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The vast majority of pyranopterin molybdenum enzymes catalyse oxygen atom transfer reactions between the substrate and solvent water coupled with proton and electron transfer.¹ The dimethyl sulfoxide reductase (DMSOR) family of pyranopterin molybdenum enzymes is unique in that they possess two pyranopterin ene-1,2-dithiolate ligands bound to the Mo ion.¹ The active site Mo centre is the locus of the oxygen atom transfer reactivity and can adopt desoxomolybdenum(IV) and monooxomolybdenum(VI) structures during the catalytic cycle.^1,2 Phylogenic analysis and protein sequence alignments of the metal-binding regions have been used to further subdivide DMSOR family enzymes into three types: I, II and III (Fig. 1).³ Type I enzymes possess a Cys or SeCys residue that coordinates to the molybdenum centre,⁴ while type II and type III enzymes have a metal center coordinated by an Asp and a Ser residue, respectively.^2,5

Fig. 1 The active site structures of oxidized type I, II and III enzymes.

Extensive studies of bis(ene-1,2-dithiolate)molybdenum complexes as active site structural models have been carried out by Holm and co-workers.^6–8 Specifically, these researchers have synthesised and structurally characterised a number of desoxomolybdenum(IV) complexes employing 1,2-dimethylethylene-1,2-dithiolate (S₂C₂Me₂) together with RS⁻, RSe⁻, RCOO⁻, and RO⁻ ligands (X)⁶ that are structurally related to the molybdenum(IV) active site structures of DMSOR type I, II and III enzymes.⁷ However, the corresponding monooxomolybdenum(VI) complexes, [Mo^VIO(L)(S₂C₂Me₂)₂]⁻, have proved too unstable for full characterisation due to an auto-redox reaction between the Mo^VI ion and the monodentate ligand L that results in the formation of the five-coordinate [Mo^VO(S₂C₂Me₂)₂]⁻ complex.^7,8 The only crystallographically characterised example of a [Mo^VIO(L)(ene-1,2-dithiolate)₂]⁻ complex is [Mo^VIO(OSiⁱPr₃)(S₂C₂(COOMe)₂)₂]⁻ (Mo^VIO(OSiⁱPr₃)(L^COOMe)₂, S₂C₂(COOMe)₂ = 1,2-dicarbomethoxyethylene-1,2-dithiolate), where the six-coordinate structure appears to be stabilised by the presence of electron-withdrawing methoxycarbonyl (–COOMe) groups on the dithiolene.⁹Mo^VIO(OSiⁱPr₃)(L^COOMe)₂ best represents a synthetic analogue of the DMSOR type III enzyme active sites. Recently, we prepared and characterised oxosulfido- and oxoselenido-molybdenum(VI) complexes at low temperature as active site models for the xanthine oxidase family of pyranopterin molybdenum enzymes.¹⁰ Herein, we report the successful application of low temperature techniques to generate and characterise new bis(ene-1,2-dithiolate)oxocarboxylatomolybdenum(VI) complexes, which can be regarded as structural models for oxidised members of type II DMSOR family enzymes. Additionally, we provide an initial electronic structure description of these bis(ene-1,2-dithiolate)oxocarboxylatemolybdenum(VI) complexes using DFT calculations. The designation and abbreviation of the complex structures are given in Chart S1.†

Initial attempts to prepare the oxocarboxylatomolybdenum(VI) complex Mo^VIO(p-H-OBz)L₂ (L = cyclohexene-1,2-dithiolate (S₂C₂C₄H₈)) utilised an oxo-transfer reaction from the tertiary amine oxide (Me₃NO) to the benzoatomolybdenum(IV) complex, Mo^IV(p-H-OBz)L₂, at room temperature under an inert Ar atmosphere. However, the grey coloured oxomolybdenum(V) complex [Mo^VO(S₂C₂C₄H₈)₂]⁻ (Mo^VOL₂) was obtained instead of the anticipated oxocarboxylatomolybdenum(VI) complex Mo^VIO(p-H-OBz)L₂. The formation of Mo^VO(ene-1,2-dithiolate)₂ has also been reported in the reaction of [Mo^IV(X)(S₂C₂Me₂)₂]⁻ with tertiary amine oxides or sulfoxides.⁸ We attempted to observe the formation of (Mo^VIO(p-H-OBz)L₂ at low temperatures, but the reaction between Mo^IV(p-H-OBz)L₂ and tertiary amine oxides does not proceed to a significant extent below −40 °C. Next, the reaction of the five-coordinate oxomolybdenum(VI) complex, Mo^VIOL₂ (synthesis in ESI†),¹¹ with the benzoate anion, Et₄N(p-H-OBz), was examined in C₂H₅CN under an Ar atmosphere. At room temperature, this reaction also yielded the oxomolybdenum(V) complex (Mo^VOL₂) (Fig. S1†), but at low temperature (−60 °C) the reaction yielded a deep-green EPR silent product. Fig. 2 shows the observed spectral changes upon addition of p-H-OBz⁻ to Mo^VIOL₂ in C₂H₅CN at −60 °C. Here, the absorption band at 395 nm due to Mo^VIOL₂ decreases with the concomitant appearance of new absorption bands at 807 nm (band 1) and 597 nm (band 2). Tight isosbestic points are observed at 336, 367 and 445 nm. Titration plots clearly indicate that the stoichiometry of Mo^VIOL to p-H-OBz⁻ is 1 : 1 (Fig. 2, inset). The final spectrum is very similar to that of [Mo^VIO(OSiⁱPr₃)(S₂C₂(COOMe)₂)₂]⁻,⁹ consistent with the formation of a six-coordinate oxocarboxylatomolybdenum(VI) complex, [Mo^VIO(p-H-OBz)(S₂C₂C₄H₈)₂]⁻, (Mo^VIO(p-H-OBz)L₂). Mo^VIO(p-OMe-OBz)L₂ and Mo^VIO(p-Cl-OBz)L₂ possess similar spectral features to Mo^VIO(p-H-OBz)L₂ and were generated in a similar manner using Et₄N(p-OMe-OBz) and Et₄N(p-Cl-OBz), respectively (Fig. S2†). The λ_max for band 1 shifts to lower energy as the substituent X changes from electron withdrawing to electron donating (X = Cl, 793 nm; X = H, 807 nm; X = OMe, 814 nm). This strongly supports the direct coordination of the benzoate ligand to the molybdenum(VI) centre. When warmed to room temperature, the UV-vis and EPR spectra converted to those of Mo^VOL₂.

Fig. 2 Spectral changes observed upon addition of 1/5, 2/5, 3/5, 4/5, and 5/5 equiv. of Et₄N(p-H-OBz) to Mo^VIOL₂ (0.2 mM) in C₂H₅CN at −60 °C; (inset) plots of the absorbance at 395 (●) and 807 (■) nm against the molar ratio of [p-H-OBz⁻]/[Mo^VIOL₂].

Cyclic voltammograms of 0.1 M ⁿBu₄NPF₆–C₂H₅CN solutions containing Mo^VIOL₂ and Et₄N(p-X-OBz) (X = Cl, H, OMe) in a 1 : 1 ratio were measured at −60 °C and yielded one irreversible reduction wave below −1 V vs. SCE. The reduction peak potential is observed to shift in a negative direction (Cl, E_pc = −1.08 V; H, E_pc = −1.16; OMe, E_pc = −1.18 V vs. SCE) as the pK_a value of the p-substituted benzoic acid increases (3.99 for X = Cl, 4.20 for X = H and 4.50 for X = OMe). This result provides support for the coordination of the p-X-OBz anion to the Mo center of Mo^VIOL₂ since the electron-donating substituent increases the basicity of the benzoate anion and enhances its ability to coordinate to Mo^VI. The CSI-mass spectrum of a C₂H₅CN solution containing Mo^VIOL₂ and 1 equiv. of Et₄N(p-OMe-OBz) showed a peak cluster attributable to [Mo(p-OMe-OBz)(S₂C₂C₄H₈)₂]⁻ at m/z = 537 at −60 °C (Fig. S3†). Since the UV-vis spectrum of the C₂H₅CN solution of the product shown in Fig. 2 is different from that of [Mo^IV(p-OMe-OBz)(S₂C₂C₄H₈)₂]⁻ synthesised separately from [Mo^IV(S₂C₂C₄H₈)₂] and p-OMe-OBz⁻, this peak cluster is likely to be a fragment of Mo^VIO(p-OMe-OBz)L₂. Therefore, we conclude that Mo^VIO(p-H-OBz)L₂ and its para substituted derivatives are formed at low temperature by coordination of the benzoates to the molybdenum(VI) centre of Mo^VIOL₂.

In order to obtain information about the mechanism of Mo^VIO(p-X-OBz)L₂ formation, a low temperature kinetic study was performed. The spectral changes observed upon addition of the benzoate ligand to Mo^VIOL₂ were observed to be biphasic. The data show a rapid disappearance of the Mo^VIOL₂ 395 nm band and the appearance of a new absorption band at 360 nm due to intermediate A that converts to Mo^VIO(p-X-OBz)L₂ with characteristic absorption bands at 597 and 807 nm (Fig. S4a and 4b†). Although the time course of the first step was too fast to be followed accurately, the conversion of intermediate A to Mo^VIO(p-X-OBz)L₂ obeys first-order kinetics. It should be noted that the observed first-order rate constant, k_obs, was independent of the concentration of Et₄N(p-H-OBz) (Fig. S5†). The formation of Mo^VIO(p-Cl-OBz)L₂ and Mo^VIO(p-OMe-OBz)L₂ displayed kinetic behavior similar to Mo^VIO(p-H-OBz)L₂ with k_obs increasing as the electron-withdrawing character of X increases: 65.8 × 10⁻³ s⁻¹ for X = Cl, 7.2 × 10⁻³ s⁻¹ for X = H and 1.4 × 10⁻³ s⁻¹ for X = OMe. These observations suggest that the first step is the association (coordination) of the benzoate anion with the Mo center of Mo^VIOL₂, giving intermediate A, and the second step is the intramolecular rearrangement of intermediate A to the product (Scheme 1). DFT calculations support an idealised C_2v structure with respect to the two S₂C₂C₄H₈ ligands for intermediate A that subsequently rearranges via a Ray–Dutt type twist¹² to form a more stable product with a distorted octahedral geometry (Fig. 3).

Scheme 1 Proposed mechanism for formation of Mo^VIO(p-X-OBz)L.

Fig. 3 Computed structures of intermediate A (left) and Mo^VIO(p-H-OBz)L₂ (right).

The DFT optimised structure of Mo^VIO(p-H-OBz)L₂ possesses a distorted octahedral coordination environment that is similar to the related Mo^VIO(OSiⁱPr₃)(L^COOMe)₂ (vide supra; Fig. 3), which has been previously characterised by X-ray crystallography.⁹ The computations indicate a slightly larger S1–S2–S3–S4 dithiolene dihedral angle for Mo^VIO(p-H-OBz)L₂ of 126° compared with a 108° dihedral angle for structurally characterised Mo^VIO(OSiⁱPr₃)(L^COOMe)₂. As was observed for Mo^VIO(OSiⁱPr₃)(L^COOMe)₂, the Mo–S4 bond distance (2.55 Å) is elongated when compared to the three Mo–S bonds (mean 2.42 Å) as a result of a strong trans influence due to the Mo≡O bond. Geometry optimisations indicate that Mo^VIO(p-Cl-OBz)L₂ and Mo^VIO(p-OMe-OBz)L₂ possess S2–S1–S3–S4 dihedral angles, and Mo–S, Mo≡O_oxo and Mo–O_OBz bond distances, which are nearly identical to those of Mo^VIO(p-H-OBz)L₂. The computed structures for the Mo^VIO(p-X-OBz)L₂ complexes display ∼172° O_oxo–Mo–S3–C dihedral angles, allowing for strong Mo d_xy–S_dithiolene π-bonding involving a single S3 donor on dithiolene B. This is supported by bonding calculations that show a LUMO wavefunction which possesses a strong Mo–S_dithiolene π* bonding interaction between the Mo(d_xy) orbital and the cis S3(p_z) donor orbital located on dithiolene (B) (Table 1). This Mo–S π* bonding description is similar to what we observed in the related complex Mo^VIO(OSiⁱPr₃)(L^COOMe)₂,⁹ and this derives from the ∼180° O_oxo–Mo–S–C dihedral angle involving the cis S of dithiolene B.

Table 1 Molecular orbital compositions for Mo^VIO(p-H-OBz)L₂

Molecular orbital	Fragment character (%)
	Mo	Oxo	OBz	S2C4H8A^a	S2C4H8B^a
a Values in parentheses are the S contribution of the dithiolene.
LUMO + 2	5	0	91	1 (1)	1 (1)
LUMO + 1	59	22	5	8 (4)	6 (5)
LUMO	41	1	11	10 (9)	38 (29)
HOMO	5	5	1	63 (37)	26 (18)
HOMO − 1	18	4	4	24 (12)	50 (26)

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The electronic absorption spectra of the Mo^VIO(p-X-OBz)L₂ complexes display well-resolved bands at ∼800 nm and ∼600 nm (band 1: ∼12 500 cm⁻¹, ε ∼ 1375 M⁻¹ cm⁻¹; and band 2: 16 700 cm⁻¹, ε ∼ 1600 M⁻¹ cm⁻¹, respectively). Using a combination of time-dependent DFT computations and our previous band assignments for Mo^VIO(OSi^iPr3)(^LCOOMe)₂,⁹ we assigned band 1 as dominantly deriving from a HOMO → LUMO one-electron promotion with appreciable dithiolene A → (Mo d_xy + dithiolene B) charge transfer character. The dominant contributor to band 2 is a HOMO − 1 → LUMO (Mo–S3_dithiolene π → Mo–S3_dithiolene π*) one-electron promotion with dithiolene A → Mo(d_xy) charge transfer character. The computations also suggest a variable HOMO → LUMO + 2 contribution with benzoate acceptor character that leads to intensity differences in band 2.

Conclusions

New bis(ene-1,2-dithiolato)oxocarboxylatomolybdenum(VI) complexes have been synthesized as active site analogues of type II DMSOR family enzymes. Low temperature kinetic and spectroscopic analyses, in conjunction with DFT calculations, have been used to understand their formation and electronic structure. Provided DMSOR_ox possesses a hexacoordinate geometry; these results suggest that the ancillary O_asp ligand likely functions to fine tune the redox potential of the Mo ion. Interestingly, a hexacoordinate geometry also allows for a specific pyranopterin dithiolene to couple the active site into long-range superexchange pathways for electron transfer regeneration of the catalytically relevant Mo(IV) site.

This work was partly supported by grants (no. 23350027, 24108725 and 2410915 to H.S. and no. 22105007 to S.I.). M.L.K. acknowledges the National Institutes of Health (GM 057378) for financial support. The authors also thank Dr Kei Ohkubo and Prof. Shunichi Fukuzumi of Osaka University for their help in collecting the EPR spectrum.

Notes and references

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Footnote

† Electronic supplementary information (ESI) available: Experimental details for the preparation, low temperature measurements and DFT calculations (Tables S1–S2, Fig. S1–S8). See DOI: 10.1039/c3dt51485d

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