The mechanism of action of ethanolamine ammonia-lyase, an adenosylcobalamin-dependent enzyme. Evidence that the hydrogen transfer mechanism involves a second intermediate hydrogen carrier in addition to the cofactor.

Tritium isotope effects in adenosylcobalamin-dependent glutamate mutase: Implications for the mechanism

“…Possibility of Tritium Transfer to a Protein-Based Intermediate. Tritium transfer to a labile site on the protein has been demonstrated for AdoCbl-dependent ribonucleotide reductase (Hogenkamp et al, 1968) and ethanolamine ammonia-lyase (O'Brien et al, 1985). It was therefore of interest to determine whether tritium is transferred to the protein during the course of the glutamate mutase reaction.…”

“…The protein radical is thought to reside on Cys-408, based on mutagenesis experiments (Booker et al, 1994) and sequence comparisons with the iron-dependent ribonucleotide reductase whose structure has recently been determined (Uhlin & Eklund, 1994). The very large tritium isotope effects measured for ethanolamine ammonia-lyase and diol dehydrase together with the exchange of tritium onto a labile site on the protein in ethanolamine ammonia-lyase (O'Brien et al, 1985) have been interpreted as evidence for a protein radical in these enzymes.…”

“…for the transfer of tritium between AdoCbl and product, of 160 and 125, respectively, measured for the deaminations and dehydrations catalyzed by ethanolamine ammonia-lyase and diol dehydrase (Weisblat & Babior, 1971;Essenberg et al, 1971). In the case of ethanolamine ammonia-lyase, a protein-based radical is further supported by experiments that demonstrate the release of tritium onto a labile site in the protein (O'Brien et al, 1985) and EPR studies with isotopically-labeled substrates (Tan et al, 1986). One attraction of this proposal is that it would provide a mechanistic link with AdoCbl-dependent ribonucleotide reductase in which a protein-based intermediate radical is an established feature (Booker et al, 1994).…”

Coupling of Cobalt−Carbon Bond Homolysis and Hydrogen Atom Abstraction in Adenosylcobalamin-Dependent Glutamate Mutase

“…Indeed, there is good evidence for extensive tunneling both hydrogen and deuterium in the lipoxygenase-catalyzed reaction (22). The very large tritium isotope effects, of 125 and 150, measured for AdoCbldependent diol dehydratase and ethanolamine ammonialyase, respectively (24,25), may also arise from quantum tunneling, although alternative explanations involving partioning of tritium between the substrate and a protein-based radical have been proposed (26,27). As quantum tunneling is detected in more enzymes (28), this phenomenon may prove to be an important feature of AdoCbl-mediated catalysis and possibly of radical-requiring enzymes in general.…”

Identification of a Rearranged-Substrate, Product Radical Intermediate and the Contribution of a Product Radical Trap in Vitamin B12 Coenzyme-Dependent Ethanolamine Deaminase Catalysis

“…The 5‘-deoxyadenosyl radical has been proposed 1,3,6 to abstract a hydrogen atom from substrate aminoethanol 1 in the first hydrogen transfer step, HT1, generating the 2-aminoethanol-1-yl substrate radical 2 . The additional participation of a protein-associated radical intermediate between the deoxyadenosyl and substrate-derived radicals has been proposed . The electron-deficient substrate radical 2 is thus activated for rearrangement to a product radical 3 .…”