Ternary complexes of transketolase with thiamin - PP and Mn 2 ion will be studied, first by EPR and then by high resolution (270 MHz) NMR and spin-echo NMR, to determine the nature of the binding, in an attempt to elucidate the basis of the quasi-irreversibility of the coenzyme binding. The identity or non-identity of the Mn 2 ion and Mg 2 ion binding sites will be determined. In parallel with studies of the paramagnetic Mn 2 ion, a study will be begun of the paramagnetic species VO 2 ion. Its possible role as a ligand acceptor for thiamin-PP, ATP and related nucleotides, will be investigated by the same NMR and EPR techniques. The binding of VO 2 ion to transketolase will be investigated, as well as its ability to substitute for divalent metallic cations in enzymatic catalysis. A conformational study of thiamin and some of its derivatives will be carried out by 13C-NMR; both high resolution and relaxation techniques will be used. Analogues of thiamin will be investigated to help elucidate the catalytic mechanism. BIBLIOGRAPHIC REFERENCES: 13C-NMR Studies of DL-2-(alpha-Hydroxyethyl)thiamin. A.A. Gallo and H.Z. Sable, J. Biol. Chem. 251, 2564 (1976). Structural and Mechanistic Aspects of Catalysis by Thiamin. A.A. Gallo, J.J. Mieyal and H.Z. Sable, in Bioorganic Chemistry, E.E. Van Tamelen, Ed., Vol. IV. Academic Press N.Y. (1977), in press.
NIH ExPorter
NIH ExPorter