X-ray crystallographic studies [Ogata et al., J. Am. Chem. Soc. 124 (2002) 11628-11635] have shown that carbon monoxide binds to the nickel ion at the active site of the [NiFe] hydrogenase from Desulfovibriovulgaris Miyazaki F and inhibits its catalytic function. In the present work spectroscopic aspects of the CO inhibition for this bacterial organism are reported for the first time and enable a direct comparison with the existing crystallographic data. The binding affinity of each specific redox state for CO is probed by FTIR spectro-electrochemistry. It is shown that only the physiological state Ni-SI(a) reacts with CO. The CO-inhibited product state is EPR-silent (Ni2+) and exists in two forms, Ni-SCO and Ni-SCO(red). At very negative potentials, the exogenous CO is electrochemically detached from the active site and the active Ni-R states are obtained. At temperatures below 100 K, photodissociation of the extrinsic CO from the Ni-SCO state results in Ni-SI(a) that is identified to be the only light-induced state. In the dark, rebinding of CO takes place; the recombination rate constants are of biexponential character and the activation barrier is determined to be approximately 9 kJ mol(-1). In addition, formation of a paramagnetic CO-inhibited state (Ni-CO) was observed that results from the interaction of carbon monoxide with the Ni-L state. It is proposed that the nickel in Ni-CO is in a formal monovalent state (Ni1+).
The [NiFe] hydrogenase from the sulphate-reducing bacterium Desulfovibrio vulgaris Miyazaki F is reversibly inhibited in the presence of molecular oxygen. A key intermediate in the reactivation process, Ni-SIr, provides the link between fully oxidized (Ni-A, Ni-B) and active (Ni-SIa, Ni-C and Ni-R) forms of hydrogenase. In this work Ni-SIr was found to be light-sensitive (T ≤ 110 K), similar to the active Ni-C and the CO-inhibited states. Transition to the final photoproduct state (Ni-SL) was shown to involve an additional transient light-induced state (Ni-SI1961). Rapid scan kinetic infrared measurements provided activation energies for the transition from Ni-SL to Ni-SIr in protonated as well as in deuterated samples. The inhibitor CO was found not to react with the active site of the Ni-SL state. The wavelength dependence of the Ni-SIr photoconversion was examined in the range between 410 and 680 nm. Light-induced effects were associated with a nickel-centred electronic transition, possibly involving a change in the spin state of nickel (Ni2+). In addition, at T ≤ 40 K the CN− stretching vibrations of Ni-SL were found to be dependent on the colour of the monochromatic light used to irradiate the species, suggesting a change in the interaction of the hydrogen-bonding network of the surrounding amino acids. A possible mechanism for the photochemical process, involving displacement of the oxygen-based ligand, is discussed.Electronic supplementary materialThe online version of this article (doi:10.1007/s00775-009-0566-9) contains supplementary material, which is available to authorized users.
The light-induced Ni-C to Ni-L transition results in the dissociation of a hydrogenic species, originating from the dihydrogen splitting at the active site. Back conversion in the dark to form Ni-C was investigated by studying the rebinding kinetics of this ligand in protonated (H(2)/H(2)O) and deuterated (D(2)/D(2)O) samples using time resolved FTIR spectroscopy.
SynopsisThe binding of tris(2,2'-bipyridyl)ruthenium(II) cations [Ru(bpy)$+ ] with single-and doublestranded (ss and ds) DNA, and the polynucleotides poly(A), poly(C), poly(G), poly(I), poly(1) .poly(C), and poly(U), was studied in aqueous solution. Steady-state electrical conductivity measurements with the polynucleotides, ssDNA, and dsDNA reveal that approximately three nucleotides offer one binding site. This may be compared with the ratio [nucleotide]/[Mg*+] of 2.4 : 1 for dsDNA. After laser excitation (353 nm), the luminescence of Ru(bpy)$+ bound to nucleic acids shows two decay components. The contribution of the fast component, which is interpreted as resulting from quenching processes of the absorbed ruthenium complex, exhibits a maximum with increasing [nucleotide]/[Ru(bpy)$+] at a ratio of about three to one. Bound Ru(bpy)E' can be released from the strand by addition of NaC10, [half-concentration: 2.5 and 2 10 m M for poly(U) and dsDNA, respectively].
The trans-»eis photoisomerization of a series of substituted stilbazolium salts (A t + X -, A t + : trans-l-alkyl-4-[4-R-styryl]-pyridinium and -quinolinium, R = CN. H, CH 3 and OCH 3 , X"=T and CIO4) was studied by laser flash photolysis and steady state irradiation measurements. The quantum yields of eis ^ trans photoisomerization (<£ c _ t and $ t _ c ) and of fluorescence of the trans isomers (> f ) were determined in several solvents at room temperature and at low temperatures in mixtures of either 2-methyltetrahydrofuran-dichloromethane or ethanolmethanol (E-M). In polar solvents at room temperature <£ t _ c is substantial (S 0.3) and > f is small (10 _3 -10~2). Competition of fluorescence and an activated step in the trans-»eis pathway is indicated by the effects of temperature on (f) { and > t _ c (activation energy: 2-3 kcal/mol). A transient, observed only at low temperatures (lifetime i T > 0.5 ms in E-M below -170°C), is assigned to the lowest triplet state with trans configuration. On the basis of the effects of temperature on (/> f , > t _> c , r T , and the triplet yield and those of quenchers on > f and > t _ c , involvement of the triplet state in the twisting process at room temperature is excluded. Therefore, a singlet mechanism is suggested for the trans-»•cis photoisomerization of the stilbazolium salts examined. Significant reduction of > t _ c for iodides in solvents of moderate polarity, where ion pairs are present, is accounted for by photoinduced electron transfer in competition to trans -»• cis photoisomerization.
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