Binding Subsites In the Active Site of $Zn^{2+}$-Glycerophosphocholine Cholinephosphodiesterase

  • Received : 1994.09.08
  • Published : 1995.03.31

Abstract

The properties of binding sites in the active site of $Zn^{2+}$-glycerophosphocholine cholinephosphodiesterase were examined using substrates and inhibitors of the enzyme. Phosphodiesterase hydrolyzed p-nitrophenylphosphocholine, p-aminophenylphosphocholine, and glycerophosphocholine, but did not hydrolyze either acylated glycerophosphocholine or bis (p-nitrophenyl)phosphate, suggesting a size limitation for interaction with a glyceryl moiety-binding subsite. The hydrolysis of p-nitrophenylphosphocholine was competitively inhibited by glycerophosphocholine and p-aminophenylphosphocholine, while glycerophosphoethanolamine was a weak inhibitor. The enzyme was also inhibited by choline, but not by ethanolamine. Thiocholine, a much more potent inhibitor than choline, was more inhibitory than cysteamine, suggesting a strict specificity of an anionic subsite adjacent to a $Zn^{2+}$ subsite. Of all oxyanions tested, the tellurite ion was found to strongly inhibit the enzyme by binding to a $Zn^{2+}$ subsite. The inhibitory role of tellurite was synergistically enhanced by tetraalkylammonium salts, but not by glycerol. Deactivation of the enzyme by diethylpyrocarbonate was partially protected by choline, but not by glycerophosphate. It is suggested that the active site of phosphodiesterase contains three binding subsites.

References

  1. Proteines ans Enzymes Bell, J.E.;Bell, E.T.
  2. Biochemistry v.16 Cushman, D.W.;Cheung, H.S.;Sabo, E.T.;Ondetti, M.A. https://doi.org/10.1021/bi00644a014
  3. Biochem. Phamacol. v.7 Ellman, G.L.;Courtney, K.D.;Andres, V.;Featherstone, R.M. https://doi.org/10.1016/0006-2952(61)90145-9
  4. Exp. Neurol. v.109 Janzen, L.;Tourtellotte, W.W.;Kanfer, J.N. https://doi.org/10.1016/0014-4886(90)90079-8
  5. J. Neuro. Res. v.24 Kanfer, J.N.;McCartney, D.G. https://doi.org/10.1002/jnr.490240214
  6. J. Biol. Chem. v.237 Kitz, R.;Wilson, I.B.
  7. Biochim. Biophys. Acta v.1071 Lennarz, W.J.;Strittmatter, W.J. https://doi.org/10.1016/0304-4157(91)90022-O
  8. Biochem. Biophys. Res. Comm. v.182 Martinez, A.;Olafsdottir, S.;Haavik, J.;Flatmark, T. https://doi.org/10.1016/S0006-291X(05)80116-0
  9. J. Biol. Chem. v.266 Oming, L.;Krivi, G.;Gierse, J.;Aykent, S.;Fitzpatrick, F.A.
  10. Biochem. J. v.286 Sok, D.E.;Kim, M.R. https://doi.org/10.1042/bj2860435
  11. Biochem. J. v.284 Sok. D.E.;Kim, M.R. https://doi.org/10.1042/bj2840641
  12. Biochem. J. v.301 Sok, D.E.;Kim, Y.B.;Jung, C.H.;Choi, S.C.;Cha, S.H. https://doi.org/10.1042/bj3010713