Inhibition of [FeFe]-hydrogenase by formaldehyde: proposed mechanism and reactivity of FeFe alkyl complexes

The mechanism for inhibition of [FeFe]-hydrogenases by formaldehyde is examined with model complexes. Key findings: (i) CH2 donated by formaldehyde covalently link Fe and the amine cofactor, blocking the active site and (ii) the resulting Fe-alkyl is a versatile electrophilic alkylating agent. Solutions of Fe2[(μ-SCH2)2NH](CO)4(PMe3)2 (1) react with a mixture of HBF4 and CH2O to give three isomers of [Fe2[(μ-SCH2)2NCH2](CO)4(PMe3)2]+ ([2]+). X-ray crystallography verified the NCH2Fe linkage to an octahedral Fe(ii) site. Although [2]+ is stereochemically rigid on the NMR timescale, spin-saturation transfer experiments implicate reversible dissociation of the Fe–CH2 bond, allowing interchange of all three diastereoisomers. Using 13CH2O, the methylenation begins with formation of [Fe2[(μ-SCH2)2N13CH2OH](CO)4(PMe3)2]+. Protonation converts this hydroxymethyl derivative to [2]+, concomitant with 13C-labelling of all three methylene groups. The Fe–CH2N bond in [2]+ is electrophilic: PPh3, hydroxide, and hydride give, respectively, the phosphonium [Fe2[(μ-SCH2)2NCH2PPh3](CO)4(PMe3)2]+, 1, and the methylamine Fe2[(μ-SCH2)2NCH3](CO)4(PMe3)2. The reaction of [Fe2[(μ-SCH2)2NH](CN)2(CO)4]2− with CH2O/HBF4 gave [Fe2[(μ-SCH2)2NCH2CN](CN)(CO)5]− ([4]−), the result of reductive elimination from [Fe2[(μ-SCH2)2NCH2](CN)2(CO)4]−. The phosphine derivative [Fe2[(μ-SCH2)2NCH2CN](CN)(CO)4(PPh3)]− ([5]−) was characterized crystallographically.


Exchange Process in [2]BAr F 4 Determined by Spin Saturation Transfer
1D Spin-Saturation Transfer (SST) experiment ( both 31 P and 1 H NMR) was used to monitor the slow exchange process in [2]BAr F 4 in CD2Cl2 at 298 K. One signal is selectively irradiated, and the signal intensity of the other peak connected with the irradiated signal via chemical exchange is monitored.
For exchange process of Selectively irradiation of peak A, the intensity of peak B changes following the equation below, and the exchange rate can be extracted: Where t is the irradiation time applied to peak A, T1 is the longitudinal (spin-lattice) relaxation time of peak B (measured by inversion-recovery method here), τ is the life time of nuclei stays at site B, and the exchange rate kB = 1/τ. S16 Figure S13. Stacked           According to the IR spectra, no reaction is evident when 1 was treated with PhCHO. In the presence of HBF 4 4 ) was formed as the protonation product.  Figure S34. 1 [5]) in CD2Cl2 at room temperature. S38 Figure S35. 31 [5]) in CD2Cl2 at room temperature. S39 Figure S36. 13 [5]) in CD2Cl2 at room temperature.   with a multi-layer mirror monochromator provided the incident beam. The sample was mounted on a 0.3 mm nylon loop with the minimal amount of Paratone-N oil. Data was collected as a series of φ and/or ω scans. Data was collected at 100 K using a cold stream of N2(g). The collection, cell refinement, and integration of intensity data was carried out with the APEX3 software. 1 A multi-scan absorption correction was performed with SADABS 2 . The structure was phased with intrinsic phasing methods using SHELXT 3 and refined with the full-matrix least-squares program SHELXL. 4 A structural model consisting of three target molecules, three BF4 anions, and two dichloromethane solvent molecules in the asymmetric unit was developed.

, ([Fe 2 [(SCH 2 ) 2 NH 2 ](CO) 4 (PMe 3 ) 2 ]BF 4 ) ([1H]BF
Two of the three BF4 anions were modeled as disordered over two orientations. Within each anion, all 1,2 and 1,3 distances were restrained to be similar (esd 0.01, 0.02 Å). Rigid-bond restraints (esd 0.004) were imposed on displacement parameters for all disordered sites and similar displacement amplitudes (esd 0.005) were imposed on disordered sites overlapping by less than the sum of van der Waals radii. The site occupancy ratios were allowed to freely refine.
Both dichloromethane solvent molecules were modeled as disordered over two orientations. Within each solvent molecule, all 1,2 and 1,3 distances were restrained to be similar (esd 0.01, 0.02 Å). Similar displacement amplitudes (esd 0.005) were imposed on disordered sites overlapping by less than the sum of van der Waals radii. The site occupancy ratios were allowed to freely refine.
H atom treatment -Methyl H atom positions, R-CH3, were optimized by rotation about R-C bonds with idealized C-H, R--H and H--H distances. All of the amine H atom positions were located in the difference map; their positions were allowed to freely refine. At convergence, all amine H atoms were in good H-bonding geometries. Remaining H atoms were included as riding idealized contributors. Methyl and amine H atom U's were assigned as 1.5 times Ueq of the carrier atom; remaining H atom U's were assigned as 1.2 times carrier Ueq.
The -3 5 0 reflection was omitted from the final refinement due to being partially obscured by the beam stop support in some orientations.

Ed15Ls
([6]BF4) CCDC: 2104105 Intensity data were collected on a Bruker D8 Venture kappa diffractometer equipped with a Photon-II CPAD detector. An Iµs microfocus Mo source (λ = 0.71073 Å) coupled with a multi-layer mirror monochromator provided the incident beam. The sample was mounted on a 0.3 mm nylon loop with the minimal amount of Paratone-N oil. Data was collected as a series of φ and/or ω scans. Data was collected at 100 K using a cold stream of N2(g). The collection, cell refinement, and integration of intensity data was carried out with the APEX3 software. 1 A multi-scan absorption correction was performed with SADABS 2 . The structure was phased with intrinsic phasing methods using SHELXT 3 and refined with the full-matrix least-squares program SHELXL. 4 A structural model consisting of the target molecule, a BF4 counter ion, and a disordered solvent molecule position in the asymmetric unit was developed.
The solvent molecule position was refined as occupational disorder of diethyl ether and dichloromethane. The dichloromethane occupancy was only approximately seven percent; to maintain a reasonable geometry with such a low occupancy it was refined as an idealized, rigid fragment. 5 Similar displacement amplitudes (esd 0.01) were imposed on disordered sites overlapping by less than the sum of van der Waals radii. The site occupancy ratio was allowed to freely refine. The 1 1 0, 1 0 0, and 0 1 1 reflections were omitted from the final refinement due to being partially obscured by the beam stop in some orientations. The 6 6 12 reflection was omitted from the final refinement due to being partially obscured by the Cu beam stop in some orientations. The -8 1 3 and -9 1 2 reflections both showed large Fo 2 vs. Fc 2 deviations with Fo 2 being larger than Fc 2 . Inspection of individual frame images revealed that in several instances there was a hot pixel on the detector in close enough proximity to the reflection that it may have been included in the integration box. These reflections were omitted from the final refinement.

([2]BAr F 4) CCDC: 2104106
Intensity data were collected on a Bruker D8 Venture kappa diffractometer equipped with a Photon-II CPAD detector. An Iµs microfocus Mo source (λ = 0.71073 Å) coupled with a multi-layer mirror monochromator provided the incident beam. The sample was mounted on a 0.3 mm nylon loop with the minimal amount of Paratone-N oil. Data was collected as a series of φ and/or ω scans. Data was collected at 100 K using a cold stream of N2(g). The collection, cell refinement, and integration of intensity data was carried out with the APEX3 software. 1 A multi-scan absorption correction was performed with SADABS 2 . The structure was phased with intrinsic phasing methods using SHELXT 3 and refined with the full-matrix least-squares program SHELXL. 4 A structural model consisting of the target molecule plus one BArF24 anion in the asymmetric unit was developed.
Two of the CF3 groups on the BArF24 anion were modeled as disordered; the C20 group over two orientations and the C29 group over three orientations. Similarity restraints (esd 0.01 Å) were imposed on all disordered C---F bond distances. Within each disordered group, the fluorine displacement parameters were constrained to be the same. The C20 site occupancy ratio was allowed to freely refine. The site occupancies for the three C29 orientations were allowed to freely refine with in the restraint that the total occupancy was 1.000(1 The -1 1 1 reflection was omitted from the final refinement due to being partially obscured by the beam stop in some orientations.

Ed82Ls
(Et4N [5]) CCDC: 2104107 Intensity data were collected on a Bruker D8 Venture kappa diffractometer equipped with a Photon-II CPAD detector. An Iµs microfocus Mo source (λ = 0.71073 Å) coupled with a multi-layer mirror monochromator provided the incident beam. The sample was mounted on a 0.3 mm nylon loop with the minimal amount of Paratone-N oil. Data was collected as a series of φ and/or ω scans. Data was collected at 120 K using a cold stream of N2(g). The collection, cell refinement, and integration of intensity data was carried out with the APEX3 software. 1 A multi-scan absorption correction was performed with SADABS 2 . The structure was phased with intrinsic phasing methods using SHELXT 3 and refined with the full-matrix least-squares program SHELXL. 4 A structural model consisting of the target molecule plus one tetraethylammonium cation in the asymmetric unit was developed.
The nitrile substituent on the adt bridge was modeled as disordered over two orientations. Similarity restraints (esd 0.01 Å) were imposed on all chemically equivalent bond distances. To maintain a more linear geometry about the nitrile group for the minor orientation, similarity restraints (esd 0.02 Å) were imposed on the distances between the nitrile nitrogen atoms and the CH2 H atoms. Similar displacement amplitudes were imposed on disordered sites overlapping by less than the sum of van der Waals radii. The site occupancy ratio was allowed to freely refine.
The tetraethylammonium cation was modeled as disordered over two orientations. All 1,2 and 1,3 distances of the cation were restrained to be similar (esd 0.01, 0.04 Å). Similar displacement amplitudes were imposed on disordered sites overlapping by less than the sum of van der Waals radii. The site occupancy ratio was allowed to freely refine.
H atom treatment -Methyl H atom positions, R-CH3, were optimized by rotation about R-C bonds with idealized C-H, R--H and H--H distances. Remaining H atoms were included as riding idealized contributors. Methyl H atom U's were assigned as 1.5 times Ueq of the carrier atom; remaining H atom U's were assigned as 1.2 times carrier Ueq.
The -1 1 1 and 0 2 0 reflections were omitted from the final refinement due to being partially obscured by the beam stop in some orientations.