Alkyl Radical Geometry Controls Geminate Cage Recombination in Alkylcobalamins

Lott, William B.; Chagovetz, Alexander M.; Grissom, Charles B.

doi:10.1021/ja00154a020

Cited by 55 publications

(52 citation statements)

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“…The spectral changes reported above following visible excitation of methylcobalamin are at variance with the interpretation of earlier measurements. Both Endicott and Netzel 18 and Lott et al 2 reported an absorption increase at 474 nm and inferred rapid formation of a spectrum characteristic of cob(II)alamin following excitation Figure 10. Schematic diagram illustrating the processes observed following the photolysis of methylcobalamin (a) and adenosylcobalamin (b).…”

Section: Discussionmentioning

confidence: 93%

“…[1][2][3][4][5][6] Recent studies have examined recombination following photolysis under the influence of an external magnetic field as a probe of field effects on the geminate radical pair. [1][2][3] Photolysis and recombination have also been used as probes of electronic trans-axial ligand effects and of cage effects in a protein environment. 4,5 Photoacoustic measurements have been used to determine bond dissociation energies.…”

Section: Introductionmentioning

confidence: 99%

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Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

et al. 1999

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Femtosecond to nanosecond transient absorption spectroscopy has been used to investigate the primary photochemistry of the B 12 coenzymes, methylcobalamin and 5′-deoxyadenosylcobalamin. Photolysis at excitation wavelengths in the near UV (400 nm) and visible (520-530 nm) are compared. Measurements were performed with femtosecond time resolution covering time delays of up to 9 ns. The photochemistry of methylcobalamin is found to depend strongly on excitation wavelength, while the photochemistry of adenosylcobalamin is essentially wavelength independent over the range studied. Excitation of methylcobalamin at 400 nm results in a partitioning between prompt bond homolysis and formation of a metastable cob(III)-alamin photoproduct as reported earlier [Walker, L. A., II; Jarrett, J. T.; Anderson, N. A.; Pullen, S. H.; Matthews, R. G.; Sension, R. J. J. Am. Chem. Soc. 1998, 120, 3597-3603]. Excitation of methylcobalamin at 520 nm in the visible R -band results only in formation of the metastable cob(III)alamin photoproduct. No prompt bond homolysis is observed. The metastable photoproduct partitions between formation of cob(II)-alamin (14 ( 5%) and recovery of the methylcobalamin starting material (86 ( 5%) on a 1.0 ( 0.1 ns time scale. The quantum yield for bond homolysis in methylcobalamin is determined by the wavelength-dependent partitioning between prompt homolysis and formation of the metastable photoproduct and by the partitioning of the metastable photoproduct between bond homolysis and ground-state recovery. In contrast, excitation of adenosylcobalamin at both 400 and 520 nm results in the development of a difference spectrum characteristic of the formation of cob(II)alamin on a picosecond time scale. The quantum yield for bond homolysis in this case is determined primarily by the competition between geminate recombination and diffusion to form solventseparated radical pairs. The caging fraction for adenosylcobalamin in aqueous solution at room temperature is 0.71 ( 0.05.

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Section: Discussionmentioning

confidence: 93%

Section: Introductionmentioning

confidence: 99%

Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

et al. 1999

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“…The effect of NO on methylmalonyl-CoA mutase has not been studied, and similar to methylcobalamin, NO does not react with AdoCbl (9). Analogous to the inhibition of NO for methionine synthase, it seemed possible that NO could inhibit mutase activity by binding to the Cbl(II) intermediate, or even to the free deoxyadenosyl radical intermediate because NO reacts readily with free radicals, but the two intermediates are generated only transiently, and their recombination rate back to AdoCbl is very high (18,19). Moreover, the Cbl(II) and deoxyadenosyl radical are generated at the active site, which at least in the case of the bacterial enzyme, is buried at the end of a (␤␣) 8 TIM barrel (20,21).…”

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confidence: 99%

Nitric Oxide Inhibits Mammalian Methylmalonyl-CoA Mutase

Kambo¹,

Sharma

Casteel

et al. 2005

Journal of Biological Chemistry

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Methylmalonyl-CoA mutase is a key enzyme in intermediary metabolism, and children deficient in enzyme activity have severe metabolic acidosis. We found that nitric oxide (NO) inhibits methylmalonyl-CoA mutase activity in rodent cell extracts. The inhibition of enzyme activity occurred within minutes and was not prevented by thiols, suggesting that enzyme inhibition was not occurring via NO reaction with cysteine residues to form nitrosothiol groups. Enzyme inhibition was dependent on the presence of substrate, implying that NO was reacting with cobalamin(II) (Cbl(II)) and/or the deoxyadenosyl radical (⅐CH 2 -Ado), both of which are generated from the co-factor of the enzyme, 5-deoxyadenosyl-cobalamin (AdoCbl), on substrate binding. Consistent with this hypothesis was the finding that high micromolar concentrations (>600 M) of oxygen also inhibited enzyme activity. To study the mechanism of NO reaction with AdoCbl, we simulated the enzymatic reaction by photolyzing AdoCbl, and found that even at low NO concentrations, NO reacted with both the generated Cbl(II) and ⅐CH 2 -Ado indicating that NO could effectively compete with the back formation of AdoCbl. Thus, NO inhibition of methylmalonyl-CoA mutase appeared to be from the reaction of NO with both AdoCbl intermediates (Cbl(II) and ⅐CH 2 -Ado) generated during the enzymatic reaction. The inhibition of methylmalonyl-CoA mutase by NO was likely of physiological relevance because a NO donor inhibited enzyme activity in intact cells, and scavenging NO from cells or inhibiting cellular NO synthesis increased methylmalonyl-CoA mutase activity when measured subsequently in cell extracts.In mammalian cells only two enzymes are known to require cobalamin (vitamin B 12 ) as a cofactor: methionine synthase, which uses methylcobalamin, and methylmalonyl-coenzyme A (CoA) mutase, which uses 5Ј-deoxyadenosyl-cobalamin (AdoCbl).1 During the reaction catalyzed by methionine synthase, methylcobalamin is cleaved heterolytically generating cobalamin I (Cbl(I)) and a methyl carbocation, whereas during the reaction catalyzed by methylmalonyl-CoA mutase, AdoCbl is cleaved homolytically generating cobalamin II (Cbl(II)) and a deoxyadenosyl radical (⅐CH 2 -Ado) (1, 2). The latter serves to abstract a hydrogen atom from the substrate methylmalonylCoA, and after intramolecular migration of a thioester moiety, the hydrogen atom is transferred back to generate the product succinyl-CoA. In the final step of the reaction, the deoxyadenosyl radical recombines with Cbl(II) to regenerate AdoCbl (3). It should be noted that the homolytic cleavage of AdoCbl occurs essentially only on substrate binding to the enzyme (3). Nitric oxide (NO) is produced by a variety of mammalian cell types, and has a diverse array of cellular and physiological functions including regulation of cell growth, differentiation, and apoptosis, and modulation of blood pressure, platelet aggregation, and synaptic plasticity (4 -6). At pharmacological concentrations, NO reacts with several different chemical groups, but at phys...

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“…The existence of a magnetic field effect on the rate of geminate (Ado', CbP} recombination in buffered water (q = 1) but the absence of a magnetic field effect on the AdoCbl"' CW photolysis quantum yield in buffered water clearly shows that an external magnetic field does not affect the cage escape yield from the geminate RP. Also, the geminate RP formed during the photolysis of CH3Cb1111 does not undergo recombination (57), yet a similar magnetic field effect is observed on the CW photolysis quantum yield (2). Therefore, the three-to four-fold magnetic field effect on geminate RP re-combination of ( Ado',Cbl") is not responsible for the magnetic field dependence of the CW quantum yield in viscous solutions.…”

Section: Identity Of the Alkylcobalamin Rp Responsible For Cw Magnetimentioning

confidence: 89%

The Origin of Magnetic Field Dependent Recombination in Alkylcobalamin Radical Pairs

Natarajan

Grissom

1996

Photochem & Photobiology

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Magnetic field effect studies of alkylcobalamin photolysis provide evidence for the formation of a reactive radical pair that is born in the singlet spin state. The radical pair recombination process that is responsible for the magnetic field dependence of the continuous-wave (CW) quantum yield is limited to the diffusive radical pair. Although the geminate radical pair of adenosylcob(III)alamin also undergoes magnetic field dependent recombination (A.M. Chagovetz and C. B. Grissom, J. Am. Chem. soc. 115, 12152-12157, 1993), this process does not account for the magnetic field dependence of the CW quantum yield that is only observed in viscous solvents. Glycerol and ethylene glycol increase the microviscosity of the solution and thereby increase the lifetime of the spin-correlated diffusive radical pair. This enables magnetic field dependent recombination among spin-correlated diffusive radical pairs in the solvent cage. Magnetic field dependent recombination is not observed in the presence of nonviscosigenic alcohols such as isopropanol, thereby indicating the importance of the increased microviscosity of the medium. Paramagnetic radical scavengers that trap alkyl radicals that escape the solvent cage do not diminish the magnetic field effect on the CW quantum yield, thereby ruling out radical pair recombination among randomly diffusing radical pairs, as well as excluding the involvement of solvent-derived radicals. Magnetic field dependent recombination among alkylcobalamin radical pairs has been simulated by a semi-classical model of radical pair dynamics and recombination. These calculations support the existence of a singlet radical pair precursor.

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Alkyl Radical Geometry Controls Geminate Cage Recombination in Alkylcobalamins

Cited by 55 publications

References 1 publication

Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

Nitric Oxide Inhibits Mammalian Methylmalonyl-CoA Mutase

The Origin of Magnetic Field Dependent Recombination in Alkylcobalamin Radical Pairs

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Alkyl Radical Geometry Controls Geminate Cage Recombination in Alkylcobalamins

Cited by 55 publications

References 1 publication

Time-Resolved Spectroscopic Studies of B12 Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

Time-Resolved Spectroscopic Studies of B12 Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

Nitric Oxide Inhibits Mammalian Methylmalonyl-CoA Mutase

The Origin of Magnetic Field Dependent Recombination in Alkylcobalamin Radical Pairs

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Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent

Time-Resolved Spectroscopic Studies of B₁₂ Coenzymes: The Photolysis of Methylcobalamin Is Wavelength Dependent