Identifying molecular mechanisms of catechol o-methyltransferase activity and regulation Public Deposited

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  • March 20, 2019
Creator
  • Tsao, Douglas
    • Affiliation: College of Arts and Sciences, Department of Chemistry
Abstract
  • Catechol O-methyltransferase (COMT) is a regioselective enzyme that functions by metabolizing catecholamines such as neurotransmitters and hormones via methylation at a single hydroxyl of the catechol moiety. Two cofactors are necessary in order to catalyze the reaction: S-adenosyl-L-methionine (SAM) and a divalent metal cation. Through computational docking studies, we show that a reason metal cations may be necessary for catalysis is due to its role in aligning the donating methyl group of SAM with the nucleophile of the substrate. It is intriguing that an enzyme whose sole function is to degrade catecholamines contains highly conserved residues responsible for regioselective behavior at the catechol moiety, a function in which site-specific chemistry seems unnecessary. Our barrier height calculations for two different COMT ligands suggest that deprotonating the meta- hydroxyl leads to higher rates of methylation and consequently there is evolutionary pressure for selecting residues that can dock the ligand in an ideal conformation for efficient catalysis. Our group previously identified three major haplotypes of COMT, where two single nucleotide polymorphisms (SNPs) produce synonymous changes and an additional SNP that creates a low enzymatic activity variant (Val108Met). Here we show that the allelic variant encoding for the low activity protein shows the highest translational efficiency among the haplotypes, suggesting evolutionary selection of an RNA-structure destabilizing allele to compensate for the low activity mutation present within its protein structure. We provide a mechanism whereby destabilizing alleles may facilitate translation initiation via computational modeling of each mRNA haplotype. One of several biological factors that COMT influences is pain perception because of its role in degrading the neurotransmitters. Peripherally injected serotonin has been clinically shown to induce a hyperalgesic effect. Here we report that serotonin-induced hyperalgesia may be induced by inhibition of COMT. Our kinetic assays reveal serotonin as a non-competitive inhibitor with respect to catechol substrates. From computational modeling, we observe serotonin actively competing with the methyl donor S-adenosyl-L-methionine at the active site. Binding of COMT to serotonin inhibits the methyl donor from entering the active site, thus preventing methylation of COMT substrates.
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  • In Copyright
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  • "... in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the Department of Chemistry."
Advisor
  • Dokholyan, Nikolay
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  • Chapel Hill, NC
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  • Open access
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