Active site of brain Zn2+-glycerophosphocholine cholinephosphodiesterase and regulation of enzyme activity.
Properties of active site of Zn2+-glycerophosphocholine cholinephosphodiesterase from ox brain were examined using substrates and inhibitors of the phosphodiesterase. The anionic binding site expressed a selectivity for a positively-charged group. Meanwhile, the glyceryl moiety-binding site appeared to be a narrow crevice of a limited size, excluding the entry of acylglycerophospholipids containing long acyl chains. While endogenous quaternary ammonium compounds such as phosphocholine, choline or carnitine inhibited the enzyme, divalent metal ions such as Co2+, Mn2+ or Zn2+ enhanced the activity by 1.5 to 2-folds. The pH dependence for the inhibition by phosphocholine or the hydrolysis of substrate implies the involvement of a basic amino acid residue with a pK value of 9.6-9.7, probably lysine, in the binding of phosphoryl group. In further support, the lysine modifiers such as trinitrobenzene sulfonic acid or diethylpyrocarbonate expressed some inactivation. The pH-rate profile indicates that an amino acid residue with a pK value of 10.2, presumably tyrosine, may participate as a nucleophile in the catalysis. This might be further supported by the inactivation of the enzyme by tyrosine modifiers such as tetranitromethane or HOI-generating system. Separately, the phosphodiesterase was observed to be susceptible to the action of hydrogen peroxide or peroxynitrite-generating system. From these results, it is implied that the phosphodiesterase may be affected by endogenous sources.
Sok
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College of Pharmacy, Chungnam National University, Taejon, Korea.
D000494:Allosteric Regulation; D000818:Animals; D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D001667:Binding, Competitive / Q000187:drug effects; D001921:Brain / Q000201:enzymology; D001923:Brain Chemistry; D002384:Catalysis / Q000187:drug effects; D002413:Cations, Divalent / Q000494:pharmacology; D002417:Cattle; D002794:Choline / Q000031:analogs & derivatives / Q000494:pharmacology; D003545:Cysteine / Q000737:chemistry; D004789:Enzyme Activation / Q000187:drug effects; D005990:Glycerol / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D010084:Oxidation-Reduction; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D010767:Phosphorylcholine / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D013774:Tetranitromethane / Q000737:chemistry; D015032:Zinc / Q000378:metabolism*
D002413:Cations, Divalent; D010767:Phosphorylcholine; D001120:Arginine; D010727:Phosphoric Diester Hydrolases; C061931:glycerophosphocholine cholinephosphodiesterase; D015032:Zinc; D013774:Tetranitromethane; D003545:Cysteine; D002794:Choline; D005990:Glycerol