Spectroscopic characterization of active-site variants of the PduO-type ATP:corrinoid adenosyltransferase from Lactobacillus reuteri: insights into the mechanism of four-coordinate Co(II)corrinoid formation

The PduO-type adenosine 5'-triphosphate (ATP):corrinoid adenosyltransferase from Lactobacillus reuteri (LrPduO) catalyzes the transfer of the adenosyl-group of ATP to Co(1+)cobalamin (Cbl) and Co(1+)cobinamide (Cbi) substrates to synthesize adenosylcobalamin (AdoCbl) and adenosylcobinamide (AdoCbi(+)), respectively. Previous studies revealed that to overcome the thermodynamically challenging Co(2+) → Co(1+) reduction, the enzyme drastically weakens the axial ligand-Co(2+) bond so as to generate effectively four-coordinate (4c) Co(2+)corrinoid species. To explore how LrPduO generates these unusual 4c species, we have used magnetic circular dichroism (MCD) and electron paramagnetic resonance (EPR) spectroscopic techniques. The effects of active-site amino acid substitutions on the relative yield of formation of 4c Co(2+)corrinoid species were examined by performing eight single-amino acid substitutions at seven residues that are involved in ATP-binding, an intersubunit salt bridge, and the hydrophobic region surrounding the bound corrin ring. A quantitative analysis of our MCD and EPR spectra indicates that the entire hydrophobic pocket below the corrin ring, and not just residue F112, is critical for the removal of the axial ligand from the cobalt center of the Co(2+)corrinoids. Our data also show that a higher level of coordination among several LrPduO amino acid residues is required to exclude the dimethylbenzimidazole moiety of Co(II)Cbl from the active site than to remove the water molecule from Co(II)Cbi(+). Thus, the hydrophilic interactions around and above the corrin ring are more critical to form 4c Co(II)Cbl than 4c Co(II)Cbi(+). Finally, when ATP analogues were used as cosubstrate, only "unactivated" five-coordinate (5c) Co(II)Cbl was observed, disclosing an unexpectedly large role of the ATP-induced active-site conformational changes with respect to the formation of 4c Co(II)Cbl. Collectively, our results indicate that the level of control exerted by LrPduO over the timing for the formation of the 4c Co(2+)corrinoid intermediates is even more exquisite than previously anticipated.     

Spectroscopic and computational studies on the adenosylcobalamin-dependent methylmalonyl-CoA mutase: evaluation of enzymatic contributions to Co-C bond activation in the Co3+ ground state

Methylmalonyl-CoA mutase (MMCM) is an enzyme that utilizes the adenosylcobalamin (AdoCbl) cofactor to catalyze the rearrangement of methylmalonyl-CoA to succinyl-CoA. Despite many years of dedicated research, the mechanism by which MMCM and related AdoCbl-dependent enzymes accelerate the rate for homolytic cleavage of the cofactor's Co-C bond by approximately 12 orders of magnitude while avoiding potentially harmful side reactions remains one of the greatest subjects of debate among B(12) researchers. In this study, we have employed electronic absorption (Abs) and magnetic circular dichroism (MCD) spectroscopic techniques to probe cofactor/enzyme active site interactions in the Co(3+)Cbl "ground" state for MMCM reconstituted with both the native cofactor AdoCbl and its derivative methylcobalamin (MeCbl). In both cases, Abs and MCD spectra of the free and enzyme-bound cofactor are very similar, indicating that replacement of the intramolecular base 5,6-dimethylbenzimidazole (DMB) by a histidine residue from the enzyme active site has insignificant effects on the cofactor's electronic properties. Likewise, spectral perturbations associated with substrate (analogue) binding to holo-MMCM are minor, arguing against substrate-induced enzymatic Co-C bond activation. As compared to the AdoCbl data, however, Abs and MCD spectral changes for the sterically less constrained MeCbl cofactor upon binding to MMCM and treatment of holoenzyme with substrate (analogues) are much more substantial. Analysis of these changes within the framework of time-dependent density functional theory calculations provides uniquely detailed insight into the structural distortions imposed on the cofactor as the enzyme progresses through the reaction cycle. Together, our results indicate that, although the enzyme may serve to activate the cofactor in its Co(3+)Cbl ground state to a small degree, the dominant contribution to the enzymatic Co-C bond activation presumably comes through stabilization of the Co(2+)Cbl/Ado. post-homolysis products.     

Co-C bond activation in methylmalonyl-CoA mutase by stabilization of the post-homolysis product Co2+ cobalamin

Despite decades of research, the mechanism by which coenzyme B12 (adenosylcobalamin, AdoCbl)-dependent enzymes promote homolytic cleavage of the cofactor's Co-C bond to initiate catalysis has continued to elude researchers. In this work, we utilized magnetic circular dichroism spectroscopy to explore how the electronic structure of the reduced B12 cofactor (i.e., the post-homolysis product Co2+ Cbl) is modulated by the enzyme methylmalonyl-CoA mutase. Our data reveal a fairly uniform stabilization of the Co 3d orbitals relative to the corrin pi/pi*-based molecular orbitals when Co2+ Cbl is bound to the enzyme active site, particularly in the presence of substrate. Contrastingly, our previous studies (Brooks, A. J.; Vlasie, M.; Banerjee, R.; Brunold, T. C. J. Am. Chem. Soc. 2004, 126, 8167-8180.) showed that when AdoCbl is bound to the MMCM active site, no enzymatic perturbation of the Co3+ Cbl electronic structure occurs, even in the presence of substrate (analogues). Collectively, these observations provide direct evidence that enzymatic Co-C bond activation involves stabilization of the post-homolysis product, Co2+ Cbl, rather than destabilization of the Co3+ Cbl "ground" state.     

Spectroscopic and computational studies of the ATP:corrinoid adenosyltransferase (CobA) from Salmonella enterica: insights into the mechanism of adenosylcobalamin biosynthesis

CobA from Salmonella enterica is a member of an enzymatic system responsible for the de novo biosynthesis of adenosylcobalamin (AdoCbl), catalyzing the formation of the essential Co-C bond by transferring the adenosyl group from a molecule of ATP to a transient Co(1+)corrinoid species generated in the enzyme active site. A particularly fascinating aspect of this reaction is that the flavodoxin in vivo reducing agent that serves as the electron donor to CobA possesses a reduction potential that is considerably more positive than that of the Co(2+/1+) couple of the corrinoid substrate. To explore how CobA may overcome this challenge, we have employed electronic absorption, magnetic circular dichroism, and electron paramagnetic resonance (EPR) spectroscopies to probe the interaction between Co(3+)- and Co(2+)corrinoids and the enzyme active site. Our data reveal that while Co(3+)corrinoids interact only weakly with CobA, Co(2+)corrinoids undergo partial conversion to a new paramagnetic species that can be obtained in nearly quantitative yield when CobA is preincubated with the co-substrate ATP. This "activated" species is characterized by a distinct set of ligand field transitions in the near-IR spectral region and EPR parameters that are unprecedented for Co(2+)corrinoids. Analysis of these data on the basis of qualitative spectral correlations and density functional theory computations reveals that this unique Co(2+)corrinoid species possesses an essentially square-planar Co(2+) center that lacks any significant axial bonding interactions. Possible implications of these findings for the mechanism of Co(2+) --> Co(1+) reduction employed by CobA and Co-C bond-forming enzymes in general are explored.     

Spectroscopic studies of the corrinoid/iron-sulfur protein from Moorella thermoacetica

Methyl transfer reactions are important in a number of biochemical pathways. An important class of methyltransferases uses the cobalt cofactor cobalamin, which receives a methyl group from an appropriate methyl donor protein to form an intermediate organometallic methyl-Co bond that subsequently is cleaved by a methyl acceptor. Control of the axial ligation state of cobalamin influences both the mode (i.e., homolytic vs heterolytic) and the rate of Co-C bond cleavage. Here we have studied the axial ligation of a corrinoid iron-sulfur protein (CFeSP) that plays a key role in energy generation and cell carbon synthesis by anaerobic microbes, such as methanogenic archaea and acetogenic bacteria. This protein accepts a methyl group from methyltetrahydrofolate forming Me-Co(3+)CFeSP that then donates a methyl cation (Me) from Me-Co(3+)CFeSP to a nickel site on acetyl-CoA synthase. To unambiguously establish the binding scheme of the corrinoid cofactor in the CFeSP, we have combined resonance Raman, magnetic circular dichroism, and EPR spectroscopic methods with computational chemistry. Our results clearly demonstrate that the Me-Co3+ and Co2+ states of the CFeSP have an axial water ligand like the free MeCbi+ and Co(2+)Cbi+ cofactors; however, the Co-OH2 bond length is lengthened by about 0.2 angstroms for the protein-bound cofactor. Elongation of the Co-OH2 bond of the CFeSP-bound cofactor is proposed to make the cobalt center more "Co1+-like", a requirement to facilitate heterolytic Co-C bond cleavage.     

Influence of environment on the electronic structure of Cob(III)alamins: time-resolved absorption studies of the S(1) state spectrum and dynamics

Transient absorption spectroscopy has been used to elucidate the nature of the S1 intermediate state populated following excitation of cob(III)alamin (Cbl(III)) compounds. This state is sensitive both to axial ligation and to solvent polarity. The excited-state lifetime as a function of temperature and solvent environment is used to separate the dynamic and electrostatic influence of the solvent. Two distinct types of excited states are identified, both assigned to pi3d configurations. The spectra of both types of excited states are characterized by a red absorption band (ca. 600 nm) assigned to Co 3d --> 3d or Co 3d --> corrin pi* transitions and by visible absorption bands similar to the corrin pi-->pi* transitions observed for ground state Cbl(III) compounds. The excited state observed following excitation of nonalkyl Cbl(III) compounds has an excited-state spectrum characteristic of Cbl(III) molecules with a weakened bond to the axial ligand (Type I). A similar excited-state spectrum is observed for adenosylcobalamin (AdoCbl) in water and ethylene glycol. The excited-state spectrum of methyl, ethyl, and n-propylcobalamin is characteristic of a Cbl(III) species with a sigma-donating alkyl anion ligand (Type II). This Type II excited-state spectrum is also observed for AdoCbl bound to glutamate mutase. The results are discussed in the context of theoretical calculations of Cbl(III) species reported in the literature and highlight the need for additional calculations exploring the influence of the alkyl ligand on the electronic structure of cobalamins.     

The enzymatic activation of coenzyme B12

The enzymatic "activation" of coenzyme B12 (5'-deoxyadenosylcobalamin, AdoCbl), in which homolysis of the carbon-cobalt bond of the coenzyme is catalyzed by some 10(9)- to 10(14)-fold, remains one of the outstanding problems in bioinorganic chemistry. Mechanisms which feature the enzymatic manipulation of the axial Co-N bond length have been investigated by theoretical and experimental methods. Classical mechanochemical triggering, in which steric compression of the long axial Co-N bond leads to increased upward folding of the corrin ring and stretching of the Co-C bond is found to be feasible by molecular modeling, but the strain induced in the Co-C bond seems to be too small to account for the observed catalytic power. The modeling study shows that the effect is a steric one which depends on the size of the axial nucleotide base, as substitution of imidazole (Im) for the normal 5,6-dimethylbenzimidazole (Bzm) axial base decreases the Co-C bond labilization considerably. An experimental test was thus devised using the coenzyme analog with Im in place of Bzm (Ado(Im)Cbl). Studies of the enzymatic activation of this analog by the B12-dependent ribonucleoside triphosphate reductase from Lactobacillus leichmannii coupled with studies of the non-enzymatic homolytic lability of the Co-C bond of Ado(Im)Cbl show that the enzyme is only slightly less efficient (3.8-fold, 0.8 kcal mol(-1)) at activating Ado(Im)Cbl than at activating AdoCbl itself. This suggests, in agreement with the modeling study, that mechanochemical triggering can make only a small contribution to the enzymatic activation of AdoCbl. Another possibility, electronic stabilization of the Co(II) homolysis product by compression of the axial Co-N bond, requires that enzymatic activation be sensitive to the basicity of the axial nucleotide. Preliminary studies of the enzymatic activation of a coenzyme analog with a 5-fluoroimidazole axial nucleotide suggest that the catalysis of Co-C bond homolysis may indeed be significantly slowed by the decrease in basicity.     

Assessment of the existence of hyper-long axial Co(II)-N bonds in cobinamide B(12) models by using electron paramagnetic resonance spectroscopy.

Protein control of cobalt-axial nitrogen ligand bond length has been proposed to modulate the reactivity of vitamin B(12) coenzyme during the catalytic cycle of B(12)-dependent enzymes. In particular, hyper-long Co-N bonds may favor homolytic cleavage of the trans-cobalt-carbon bond in the coenzyme. X-ray crystallographic studies point to hyper-long bonds in two B(12) holoenzymes; however, mixed redox and ligand states in the crystals thwart clear conclusions. Since EPR theory predicts an increase in Co(II) hyperfine splitting as donation from the axial N-donor ligand decreases, EPR spectroscopy could clarify the X-ray results. However, the theory is apparently undermined by the similar splitting reported for the 2-picoline (2-pic) and pyridine (py) adducts of Co(II) cobinamide (Co(II)Cbi(+)), adducts thought to have long and normal Co-N axial bond lengths, respectively. Cobinamides, with the B(12) 5,6-dimethylbenzimidazole loop removed, are excellent B(12) models. We studied Co(II)Cbi(+) adducts of unhindered 4-substituted pyridines (4-X-py's) in ethylene glycol to separate orbital size effects from Co-N axial distance effects on these splittings. The linear increase in splitting with the decrease in 4-X-py basicity found is consistent with the theoretically predicted increase in unpaired electron spin density as axial N lone pair donation to Co(II) decreases. No adduct (and hence no hyper-long Co(II)-N axial bond) was formed even by 8 M 2-pic, if the 2-pic was purified by a novel Co(III)-affinity distillation procedure designed to remove trace nitrogenous ligand impurities present in 2-pic distilled in the regular manner. Adducts formed by impurities in 2-pic and other hindered pyridines misled previous investigators into attributing results to adducts with long Co-N bonds. We find that many 2-substituted py's known to form adducts with simple synthetic Co models do not bind Co(II)Cbi(+). Thus, the equatorial corrin ring sterically impedes binding, making Co(II)Cbi(+) a highly selective binding agent for unhindered sp(2) N-donor ligands. Our results resolve the apparent conflict between EPR experiment and theory. The reported Co(II) hyperfine splitting of the enzyme-bound cofactor in five B(12) enzymes is similar to that of the relevant free cofactor. The most reasonable interpretation of this similarity is that the Co-N axial bond of the bound cofactor is not hyper-long in any of the five cases.     

Spectroscopic and electronic structure studies of protocatechuate 3,4-dioxygenase: nature of tyrosinate-Fe(III) bonds and their contribution to reactivity.

The geometric and electronic structure of the high-spin ferric active site of protocatechuate 3,4-dioxygenase (3,4-PCD) has been examined by absorption (Abs), circular dichroism (CD), magnetic CD (MCD), and variable-temperature-variable-field (VTVH) MCD spectroscopies. Density functional (DFT) and INDO/S-CI molecular orbital calculations provide complementary insight into the electronic structure of 3,4-PCD and allow an experimentally calibrated bonding scheme to be developed. Abs, CD, and MCD indicate that there are at least seven transitions below 35 000 cm(-1) which arise from tyrosinate ligand-to-metal-charge transfer (LMCT) transitions. VTVH MCD spectroscopy gives the polarizations of these LMCT bands in the principal axis system of the D-tensor, which is oriented relative to the molecular structure from the INDO/S-CI calculations. Three transitions are associated with the equatorial tyrosinate and four with the axial tyrosinate. This large number of transitions per tyrosinate is due to the pi and importantly the sigma overlap of the two tyrosinate valence orbitals with the metal d orbitals and is governed by the Fe-O-C angle and the Fe-O-C-C dihedral angles. The previously reported crystal structure indicates that the Fe-O-C angles are 133 degrees and 148 degrees for the equatorial and axial tyrosinate, respectively. Each tyrosinate has transitions at different energies with different intensities, which correlate with differences in geometry that reflect pseudo-sigma bonding to the Fe(III) and relate to reactivity. These factors reflect the metal-ligand bond strength and indicate that the axial tyrosinate-Fe(III) bond is weaker than the equatorial tyrosinate-Fe(III) bond. Furthermore, it is found that the differences in geometry, and hence electronic structure, are imposed by the protein. The consequences to catalysis are significant because the axial tyrosinate has been shown to dissociate upon substrate binding and the equatorial tyrosinate in the enzyme-substrate complex is thought to influence asymmetric binding of the chelated substrate moiety via a strong trans influence which activates the substrate for reaction with O2.     

Magnetic quantum dots: synthesis, spectroscopy, and magnetism of Co2+ - and Ni2+-doped ZnO nanocrystals

We report a method for the preparation of colloidal ZnO-diluted magnetic semiconductor quantum dots (DMS-QDs) by alkaline-activated hydrolysis and condensation of zinc acetate solutions in dimethyl sulfoxide (DMSO). Mechanistic studies reveal that Co(2+) and Ni(2+) dopants inhibit nucleation and growth of ZnO nanocrystals. In particular, dopants are quantitatively excluded from the critical nuclei but are incorporated nearly isotropically during subsequent growth of the nanocrystals. The smaller nanocrystal diameters that result upon doping are explained by the Gibbs-Thompson relationship between lattice strain and crystal solubility. We describe methods for cleaning the nanocrystal surfaces of exposed dopants and for redispersion of the final DMS-QDs. Homogeneous substitutional doping is verified by high-resolution low-temperature electronic absorption and magnetic circular dichroism (MCD) spectroscopies. A "giant Zeeman effect" is observed in the band gap transition of Co(2+):ZnO DMS-QDs. MCD and Zeeman spectroscopies are used to quantify the magnitude of the p-d exchange interaction (N(0)beta) that gives rise to this effect. N(0)beta values of -2.3 +/- 0.3 eV (-18 500 cm(-1)) for Co(2+):ZnO and -4.5 +/- 0.6 eV (-36 300 cm(-1)) for Ni(2+):ZnO have been determined. Ligand-to-metal charge-transfer transitions are observed in the MCD spectra of both Co(2+):ZnO and Ni(2+):ZnO DMS-QDs and are analyzed in the context of an optical electronegativity model. The importance of these charge-transfer states in determining N(0)beta is discussed. Ferromagnetism with T(C) > 350 K is observed in aggregated nanocrystals of Co(2+):ZnO that unambiguously demonstrates the existence of intrinsic high-T(C) ferromagnetism in this class of DMSs.     

Spectroscopic and computational studies of Co1+cobalamin: spectral and electronic properties of the "superreduced" B12 cofactor

The 4-coordinate, low-spin cob(I)alamin (Co1+Cbl) species, which can be obtained by heterolytic cleavage of the Co-C bond in methylcobalamin or the two-electron reduction of vitamin B12, is one of the most powerful nucleophiles known to date. The supernucleophilicity of Co1+Cbl has been harnessed by a number of cobalamin-dependent enzymes, such as the B12-dependent methionine synthase, and by enzymes involved in the biosynthesis of B12, including the human adenosyltransferase. The nontoxic nature of the Co1+Cbl supernucleophile also makes it an attractive target for the in situ bioremediation of halogenated waste. To gain insight into the geometric, electronic, and vibrational properties of this highly reactive species, electronic absorption, circular dichroism (CD), magnetic CD, and resonance Raman (rR) spectroscopies have been employed in conjunction with density functional theory (DFT), time-dependent DFT, and combined quantum mechanics/molecular mechanics computations. Collectively, our results indicate that the supernucleophilicity of Co1+Cbl can be attributed to the large destabilization of the Co 3dz2-based HOMO and its favorable orientation with respect to the corrin macrocycle, which minimizes steric repulsion during nucleophilic attack. An intense feature in the CD spectrum and a prominent peak in the rR spectra of Co1+Cbl have been identified that may serve as excellent probes of the nucleophilic character, and thus the reactivity, of Co1+Cbl in altered environments, including enzyme active sites. The implications of our results with respect to the enzymatic formation and reactivity of Co1+Cbl are discussed, and spectroscopic trends along the series from Co3+Cbls to Co2+Cbl and Co1+Cbl are explored.     

Solution structure and thermolysis of Cobeta-5'-deoxyadenosylimidazolylcobamide, a coenzyme B12 analogue with an imidazole axial nucleoside

The solution structure of Cobeta-5'-deoxyadenosylimidazolylcobamide, Ado(Im)Cbl, the coenzyme B(12) analogue in which the axial 5,6-dimethylbenzimidazole (Bzm) ligand is replaced by imidazole, has been determined by NMR-restrained molecular modeling. A two-state model, in which a conformation with the adenosyl moiety over the southern quadrant of the corrin and a conformation with the adenosyl ligand over the eastern quadrant of the corrin are both populated at room temperature, was required by the nOe data. A rotation profile and molecular dynamics simulations suggest that the eastern conformation is the more stable, in contrast to AdoCbl itself in which the southern conformation is preferred. Consensus structures of the two conformers show that the axial Co-N bond is slightly shorter and the corrin ring is less folded in Ado(Im)Cbl than in AdoCbl. A study of the thermolysis of Ado(Im)Cbl in aqueous solution (50-125 degrees C) revealed competing homolytic and heterolytic pathways as for AdoCbl but with heterolysis being 9-fold faster and homolysis being 3-fold slower at 100 degrees C than for AdoCbl. Determination of the pK(a)'s for the Ado(Im)Cbl base-on/base-off reaction and for the detached imidazole ribonucleoside as a function of temperature permitted correction of the homolysis and heterolysis rate constants for the temperature-dependent presence of the base-off species of Ado(Im)Cbl. Activation analysis of the resulting rate constants for the base-on species show that the entropy of activation for Ado(Im)Cbl homolysis (13.7 +/- 0.9 cal mol(-1) K(-1)) is identical with that of AdoCbl (13.5 +/- 0.7 cal mol(-1) K(-1)) but that the enthalpy of activation (34.8 kcal mol(-1)) is 1.0 +/- 0.4 kcal mol(-1) larger. The opposite effect is seen for heterolysis, where the enthalpies of activation are identical but the entropy of activation is 5 +/- 1 cal mol(-1) K(-1) less negative for Ado(Im)Cbl. Extrapolation to 37 degrees C provides a rate constant for Ado(Im)Cbl homolysis of 2.1 x 10(-9) s(-1), 4.3-fold smaller than for AdoCbl. Combined with earlier results for the enzyme-induced homolysis of Ado(Im)Cbl by the ribonucleoside triphosphate reductase from Lactobacillus leichmannii, the catalytic efficiency of the enzyme for homolysis of Ado(Im)Cbl at 37 degrees C can be calculated to be 4.0 x 10(8), 3.8-fold, or 0.8 kcal mol(-1), smaller than for AdoCbl. Thus, the bulky Bzm ligand makes at best a <1 kcal mol(-1) contribution to the enzymatic activation of coenzyme B(12).     

Spectroscopic evidence for the formation of a four-coordinate Co2+ cobalamin species upon binding to the human ATP:cobalamin adenosyltransferase

The human adenosyltransferase hATR converts exogenous cobalamin into coenzyme B12 by transferring the adenosyl group from cosubstrate ATP to a transiently formed Co1+cobalamin (Co1+Cbl) species. A particularly puzzling aspect of hATR function is that the midpoint potential for Co2+Cbl --> Co1+Cbl reduction is below that of readily available biological reductants. Our magnetic circular dichroism and electron paramagnetic resonance spectroscopic studies reported here reveal that, in the absence of ATP, the interaction between Co2+Cbl and hATR promotes partial conversion of the cofactor to its "base-off" form in which a water molecule occupies the lower axial position. This interaction becomes much stronger in the presence of ATP, leading to the formation of an unprecedented Co2+Cbl species with spectroscopic signatures consistent with an essentially four-coordinate, square-planar Co2+ center. This unusual Co2+Cbl coordination is expected to raise the Co2+/1+ reduction potential well into the physiological range.     

X-ray structures and homolysis of some alkylcobalt(III) phthalocyanine complexes

The first crystallographic data for sigma-bonded alkylcobalt(III) phthalocyanine complexes are reported. A single-crystal X-ray structure of CH(3)CH(2)Co(III)Pc (Pc = dianion of phthalocyanine) reveals that the solid consists of centrosymmetric face-to-face dimers in which the CH(3)CH(2)Co(III)Pc units retain their square pyramidal geometry. The structure appears to be the first one reported for a five-coordinate RCo(III)(chelate) complex with an electron-deficient equatorial system. The Co-C bond in CH(3)CH(2)Co(III)Pc (2.031(5) A) is the longest found in five-coordinate RCo(III)(chel) complexes (R = simple primary alkyl group). Another X-ray study demonstrates that CH(3)Co(III)Pc(py) has a distorted octahedral geometry with axial bonds of very similar length to those in methylcobalamin. The axial bonds are shorter than those in its octaethylporphyrin analogue, in accordance with a weaker trans axial influence in six-coordinate complexes containing an electron-deficient phthalocyanine equatorial ligand. A different trend has been observed for five-coordinate RCo(III)(chel) complexes: electron-rich equatorial systems seem to make the Co-C axial bond shorter. Kinetic data for the homolysis of RCo(III)Pc complexes (R = Me, Et) in dimethylacetamide are also reported. Homolysis of ethyl derivatives is faster. The Co-C bond dissociation energies (BDEs) for the pyridine adducts of the methyl and the ethyl derivative are 30 +/- 1 and 29 +/- 1 kcal/mol, respectively. The BDE for CH(3)CoPc(py) is considerably lower than that for MeCbl despite the very similar lengths of the axial bonds in the two complexes. The results of this work do not support any correlation between the Co-C bond length and the bond strength as defined by BDE.     

Ultrafast excited-state dynamics in vitamin B12 and related cob(III)alamins

Femtosecond transient IR and visible absorption spectroscopies have been employed to investigate the excited-state photophysics of vitamin B12 (cyanocobalamin, CNCbl) and the related cob(III)alamins, azidocobalamin (N3Cbl), and aquocobalamin (H2OCbl). Excitation of CNCbl, H2OCbl, or N3Cbl results in rapid formation of a short-lived excited state followed by ground-state recovery on time scales ranging from a few picoseconds to a few tens of picoseconds. The lifetime of the intermediate state is influenced by the sigma-donating ability of the axial ligand, decreasing in the order CNCbl > N3Cbl > H2OCbl, and by the polarity of the solvent, decreasing with increasing solvent polarity. The peak of the excited-state visible absorption spectrum is shifted to ca. 490 nm, and the shape of the spectrum is characteristic of weak axial ligands, similar to those observed for cob(II)alamin, base-off cobalamins, or cobinamides. Transient IR spectra of the upper CN and N3 ligands are red-shifted 20-30 cm(-1) from the ground-state frequencies, consistent with a weakened Co-upper ligand bond. These results suggest that the transient intermediate state can be attributed to a corrin ring pi to Co 3d(z2) ligand to metal charge transfer (LMCT) state. In this state bonds between the cobalt and the axial ligands are weakened and lengthened with respect to the corresponding ground states.     

Giant excitonic Zeeman splittings in colloidal Co2+ -doped ZnSe quantum dots

Colloidal Co(2+):ZnSe diluted magnetic semiconductor quantum dots (DMS-QDs) were prepared by the hot injection method and studied spectroscopically. Ligand-field electronic absorption and magnetic circular dichroism (MCD) spectra confirm homogeneous substitutional speciation of Co(2+) in the ZnSe QDs. Absorption spectra collected at various times throughout the syntheses reveal that dopants are absent from the central cores of the QDs but are incorporated at a constant concentration during nanocrystal growth. The undoped cores are associated with dopant exclusion from the ZnSe critical nuclei. Analysis of low-temperature electronic absorption and MCD spectra revealed excitonic Zeeman splitting energies (DeltaE(Zeeman)) of these Co(2+):ZnSe QDs that were substantially smaller than anticipated from bulk Co(2+):ZnSe data. This reduction in DeltaE(Zeeman) is explained quantitatively by the absence of dopants from the QD cores, where dopant-exciton overlap would be greatest. Since dopant exclusion from nucleation appears to be a general phenomenon for DMS-QDs grown by direct chemical methods, we propose that DeltaE(Zeeman) will always be smaller in colloidal DMS-QDs grown by such methods than in the corresponding bulk materials.     

The Co-CH(3) Bond in Imine/Oxime B(12) Models. Influence of the Orientation and Donor Properties of the trans Ligand As Assessed by FT-Raman Spectroscopy

Near-IR FT-Raman spectroscopy was used to probe the properties of three types of methyl imine/oxime B(12) model compounds in CHCl(3) solution. These types differ in the nature of the 1,3-propanediyl chain and were selected to test the influence of electronic and steric effects on the Co-CH(3) stretching (nu(Co)(-)(CH)()3) frequency, a parameter related to Co-C bond strength. For the first type studied, [LCo((DO)(DOH)pn)CH(3)](0/+) ((DO)(DOH)pn = N(2),N(2)(')-propane-1,3-diylbis(2,3-butanedione 2-imine 3-oxime)), nu(Co)(-)(CH)()3 decreased from 505 to 455 cm(-)(1) with stronger electron-donating character of the trans axial ligand, L, in the order Cl(-), MeImd, Me(3)Bzm, 4-Me(2)Npy, py, 3,5-Me(2)PhS(-), PMe(3), and CD(3)(-). This series thus allows the first assessment of the effect of negative axial ligands on nu(Co)(-)(CH)()3; these ligands (L = Cl(-), 3,5-Me(2)PhS(-), CD(3)(-)) span the range of trans influence. The CH(3) bending (delta(CH3)) bands were observed at 1171, 1159, and 1150/1105 cm(-)(1), respectively. The decrease in C-H stretching frequencies (nu(CH)) of the axial methyl suggests that the C-H bond strength decreases in the order Cl(-) > 3,5-Me(2)PhS(-) > CD(3)(-). This result is consistent with the order of decreasing (13)C-(1)H NMR coupling constants obtained for the axial methyl group. The trend of lower nu(Co)(-)(CH)()3 and nu(CH) frequencies and lower axial methyl C-H coupling constant for stronger electron-donating trans axial ligands can be explained by changes in the electronic character of the Co-C bond. The symmetric CH(3)-Co-CH(3) mode (nu(CH)()3(-)(Co)(-)(CH)()3) for (CH(3))(2)Co((DO)(DOH)pn) was determined to be 456 cm(-)(1) (421 cm(-)(1) for (CD(3))(2)Co((DO)(DOH)pn). The L-Co-CH(3) bending mode (delta(L)(-)(Co)(-)(CH)()3) was detected for the first time for organocobalt B(12) models; this mode, which is important for force field calculations, occurs at 194 cm(-)(1) for ClCo((DO)(DOH)pn)CH(3) and at 186 cm(-)(1) for (CH(3))(2)Co((DO)(DOH)pn. The nu(Co)(-)(CH)()3 frequencies were all lower than those reported for the corresponding cobaloxime type LCo(DH)(2)CH(3) (DH = monoanion of dimethylglyoxime) models for planar N-donor L. This relationship is attributed to a steric effect of L in [LCo((DO)(DOH)pn)CH(3)](+). The puckered 1,3-propanediyl chain in [LCo((DO)(DOH)pn)CH(3)](+) forces the planar L ligands to adopt a different orientation compared to that in the cobaloxime models. The consequent steric interaction bends the equatorial ligand toward the methyl group (butterfly bending); this distortion leads to a longer Co-C bond. In a second imine/oxime type, a pyridyl ligand is connected to the 1,3-propanediyl chain and oriented so as to minimize butterfly bending. The nu(Co)(-)(CH)()3 frequency for this new lariat model was close to that of pyCo(DH)(2)CH(3). In a third type, a bulkier 2,2-dimethyl-1,3-propanediyl group replaces the 1,3-propanediyl chain. The nu(Co)(-)(CH)()3 bands for two complexes with L = Me(3)Bzm and py were 2-5 cm(-)(1) lower in frequency than those of the corresponding [L(Co((DO)(DOH)pn)CH(3)](+) complexes. The decrease in the axial nu(Co)(-)(CH)()3 frequencies is probably due to the steric effect of the equatorial ligand. Thus, the nu(Co)(-)(CH)()3 frequency can be useful for investigating both steric and electronic influences on the Co-C bond.     

Spectroscopy of photovoltaic and photoconductive nanocrystalline Co2+-doped ZnO electrodes

Photocurrent and photoconductivity measurements have been used in combination with absorption and magnetic circular dichroism (MCD) spectroscopic measurements to elucidate the mechanism of photoinduced carrier generation in nanocrystalline Co(2+):ZnO electrodes. These experiments allowed direct observation of two broad Co(2+) charge transfer (CT) bands extending throughout the visible energy range. The lower energy CT transition is assigned as a Co(2+) --> conduction band excitation (ML(CB)CT). Sensitization of this ML(CB)CT level by (4)A(2) --> (4)T(1)(P) ligand-field excitation is concluded to be responsible for the distinctive structured photocurrent action spectrum of these electrodes at ca. 14 000 cm(-1). The higher energy CT transition is assigned as a valence band --> Co(2+) excitation (L(VB)MCT) and is found to have an internal quantum efficiency for charge separation that is approximately four times larger than that of the ML(CB)CT excitation. The different internal quantum efficiencies for the two CT excitations are related to differences in excited-state wave functions arising from configuration interaction with the 1S excitonic levels of ZnO. Whereas the ML(CB)CT excited state is best described as a localized Co(3+) + e(-)(CB) configuration, the L(VB)MCT excited state (Co(+) + h(+)(VB)) has a 4-fold greater admixture of delocalized excitonic (Co(2+) + h(+)(VB) + e(-)(CB)) character in its wave function, a conclusion supported by quantitative analysis of the CT absorption intensities. Practical factors controlling the overall photovoltaic efficiencies of the photoelectrochemical cells, including electrode conductivity and porosity, were also examined.