Methyltransferase enzymes

Photoluminescent spectra for methyltetrahydrofolate and the enzyme methyltransferase. When methyltetrahydrofolate and methyltransferase are mixed, the enzyme is no longer photoluminescent, but the photoluminescence of methyltetrahydrofolate is enhanced. (Spectra courtesy of Dave Roberts, DePauw University.)... 

The first evidence for cobalamin involvement in the conversion of methanol to methane was provided by Blaylock and Stadtman [196,216-218] with extracts of methanol-grown M. barkeri they demonstrated enzymatic formation of methylcobalamin from methanol, and subsequent reduction of methylcobalamin to methane. Later Blaylock [196] showed that conversion of methanol to methylcobalamin requires a heat-stable cofactor and at least three proteins, a 100-200 kDa Bi2-enzyme (methyltransferase), a ferredoxin, and an unidentified protein. Blaylock speculated that the role of hydrogen and ferredoxin in the conversion of methanol to methylcobalamin was in the reduction of the Bi2-protein. This work led to the proposal that methylcobalamin was the direct precursor of methane in methanogenesis from various substrates [196,218]. 

The MeCbl-based enzymes (methyltransferases) catalyze the transfer of methyl groups, and the overall mechanistic scheme requires a reversible heterolytic cleavage of the Co-Me bond. The process catalyzed by AdoCbl-based enzymes (isomerase and eliminase) proceeds through a stepwise process initiated by the homolytic cleavage of the Co-C bond. ... 

Histamine AND histamine antagonists). It is formed from histidine by the enzyme L-histidine decarboxylase. In the periphery, histamine is stored ia mast cells, basophils, cells of the gastric mucosa, and epidermal cells. In the CNS, histamine is released from nerve cells and acts as a neurotransmitter. The actions of histamine ate terrninated by methylation and subsequent oxidation via the enzymes histamine-/V-methyltransferase and monoamine oxidase. 

Modulation of second-messenger pathways is also an attractive target upon which to base novel antidepressants. Rolipram [61413-54-5] an antidepressant in the preregistration phase, enhances the effects of noradrenaline though selective inhibition of central phosphodiesterase, an enzyme which degrades cycHc adenosiae monophosphate (cAMP). Modulation of the phosphatidyl iaositol second-messenger system coupled to, for example, 5-HT,, 5-HT,3, or 5-HT2( receptors might also lead to novel antidepressants, as well as to alternatives to lithium for treatment of mania. Novel compounds such as inhibitors of A-adenosyl-methionine or central catechol-0-methyltransferase also warrant attention. 

Fig. 2. Biosynthetic pathway for epinephrine, norepinephrine, and dopamine. The enzymes cataly2ing the reaction are (1) tyrosine hydroxylase (TH), tetrahydrobiopterin and O2 are also involved (2) dopa decarboxylase (DDC) with pyridoxal phosphate (3) dopamine-P-oxidase (DBH) with ascorbate, O2 in the adrenal medulla, brain, and peripheral nerves and (4) phenethanolamine A/-methyltransferase (PNMT) with. Cadenosylmethionine in the adrenal...

Together with dopamine, adrenaline and noradrenaline belong to the endogenous catecholamines that are synthesized from the precursor amino acid tyrosine (Fig. 1). In the first biosynthetic step, tyrosine hydroxylase generates l-DOPA which is further converted to dopamine by the aromatic L-amino acid decarboxylase ( Dopa decarboxylase). Dopamine is transported from the cytosol into synaptic vesicles by a vesicular monoamine transporter. In sympathetic nerves, vesicular dopamine (3-hydroxylase generates the neurotransmitter noradrenaline. In chromaffin cells of the adrenal medulla, approximately 80% of the noradrenaline is further converted into adrenaline by the enzyme phenylethanolamine-A-methyltransferase. 

Decitabine (5-aza-deoxycytosine) is an analog of the nucleoside 2 -deoxycytidine. It is believed to exert its antineoplastic effects after phosphorylation and direct incorporation into DNA and by inhibition of the enzyme DNA methyltransferase, causing hypomethylation of DNA and cellular differentiation or apoptosis. DNA hypomethylation is achieved at concentrations below those required to significantly inhibit DNA synthesis, which may promote restoration of function to genes associated with control of cellular differentiation and proliferation. Cytotoxicity in rapidly dividing cells may also result from covalent adducts between DNA methyltransferase and decitabine. 

Catechol O-methyltransferase (COMT) is a widespread enzyme that catalyzes the transfer of the methyl group of S-adenosyl-l-methionine (AdoMet) to one of the phenolic group of the catechol substrate (Fig. 1). High COMT activity is found in the liver, kidney and gut wall... 

The metabolism of foreign compounds (xenobiotics) often takes place in two consecutive reactions, classically referred to as phases one and two. Phase I is a functionalization of the lipophilic compound that can be used to attach a conjugate in Phase II. The conjugated product is usually sufficiently water-soluble to be excretable into the urine. The most important biotransformations of Phase I are aromatic and aliphatic hydroxylations catalyzed by cytochromes P450. Other Phase I enzymes are for example epoxide hydrolases or carboxylesterases. Typical Phase II enzymes are UDP-glucuronosyltrans-ferases, sulfotransferases, N-acetyltransferases and methyltransferases e.g. thiopurin S-methyltransferase. 

In vivo azathioprine is rapidly converted into its active metabolite 6-mercaptopurine by the enzyme thiopurine methyltransferase (TPMT). The active agent inhibits IMPDH function. Furthermore, it also acts as antimetabolite of the RNA and DNA synthesis particularly in T-lymphocytes leading to cell death. Due to genetic polymorphism of TPMT, therapy may fail, thus it is currently discussed whether individual patients should be monitored before the use of azathioprine. 

Thiopurine S-methyltransferase is an enzyme which inactivates the anticancer drug 6-mercaptopurine by S-methylation. 

These drugs are thought to prolong the effect of levodopa by blocking an enzyme, catechol-O-methyltransferase (COMT), which eliminates dopamine. When given with levodopa, the COMT inhibitors increase the plasma concentrations and duration of action of levodopa... 

Barton and coworkers have shown that proteins can in fact modulate the DNA electron transfer [168]. Methyltransferases are enzymes that recognize distinct DNA sequences, e.g., 5 -G CGC-3, and effect methylation by extrading the target base cytosine ( C) completely out of the DNA duplex while the remainder of the double helix is left intact. The methyltransferase Hha 1-DNA complex is a well-characterized example, revealing that the structure of the DNA is significantly but locally distorted [169,170]. In a recent study, Raj ski et al. used DNA duplex 20 containing the M.Hha I binding site between two oxidizable 5 -GG-3 sites [168] (Fig. 20). The duplex contains a complementary strand, selectively 5 -modified with a Rh intercalator that can function as a photooxidant. Upon... 

Neural cells convert tyrosine to epinephrine and norepinephrine (Figure 31—5). While dopa is also an intermediate in the formation of melanin, different enzymes hydroxylate tyrosine in melanocytes. Dopa decarboxylase, a pyridoxai phosphate-dependent enzyme, forms dopamine. Subsequent hydroxylation by dopamine P-oxidase then forms norepinephrine. In the adrenal medulla, phenylethanolamine-A -methyltransferase uti-hzes S-adenosyhnethionine to methylate the primary amine of norepinephrine, forming epinephrine (Figure 31-5). Tyrosine is also a precursor of triiodothyronine and thyroxine (Chapter 42). 

The possibility that the initial degree of methyl-esterification might be controlled by the properties of the methyltransferase enzymes was examined partial characterisation of these enzymes in suspension-cultured cells of fiax. Pectin methyltransferases beii enzymes characteristic of the Golgi apparatus [22], microsomes were fiactionated daily for ten days from suspension-cultured flax cells and incubated in the presence of C-SAM, the universal donor of methyl groups. 

Figure 1 indicates that pectin methyltransferase (PMT) activity from freeze-thawed microsomes measured without exogenous substrate was maximal at neutral pH (6.5 to 7.5). When exogenous pectic substrates of various DE had been added, similar optimal neutral pH was observed, and the activity was slightly stimulated (1.2 to 1.8 times). A second optimal pH occured at pH 5.5, but in the presence of low methylated pectin (DE 0.1). As suggested by Lineweaver and Ballou [8] to explain the behaviour of another pectic enzyme -i.e. pectin methylesterase (PME), the mobility and the activity of PMT might be influenced by the presence of polyanionic substrates. On the other hand, the existence of several forms of pectin methyltransferase in flax microsomes might be responsible for such variations of the activity. 

Coulter C, JTG Hamilton, WC McRoberts, L Kulakov, MJ Larkin, DB Harper (1999) Halomethane bisulfite/halide ion methyltransferase, an unusual corrinoid enzyme of environmental significance isolated from an aerobic methylotroph using chloromethane as the sole carbon source. Appl Environ Microbiol 65 4301-4312. 

Because LCEC had its initial impact in neurochemical analysis, it is not, surprising that many of the early enzyme-linked electrochemical methods are of neurologically important enzymes. Many of the enzymes involved in catecholamine metabolism have been determined by electrochemical means. Phenylalanine hydroxylase activity has been determined by el trochemicaUy monitoring the conversion of tetrahydro-biopterin to dihydrobiopterin Another monooxygenase, tyrosine hydroxylase, has been determined by detecting the DOPA produced by the enzymatic reaction Formation of DOPA has also been monitored electrochemically to determine the activity of L-aromatic amino acid decarboxylase Other enzymes involved in catecholamine metabolism which have been determined electrochemically include dopamine-p-hydroxylase phenylethanolamine-N-methyltransferase and catechol-O-methyltransferase . Electrochemical detection of DOPA has also been used to determine the activity of y-glutamyltranspeptidase The cytochrome P-450 enzyme system has been studied by observing the conversion of benzene to phenol and subsequently to hydroquinone and catechol... 

Mazzafera, P., Wingsle, G., Olsson, O., Sandberg, G., S-Adenosyl-L-methionine theobromine 1-N-methyltransferase, an enzyme catalyzing the synthesis of caffeine in coffee, Phytochemistry, 37(6), 1577, 1994. (CA122 259411n)... 

Figure 1. Biosynthetic pathways for biogenic amines. In Drosophila and vertebrates decarboxylation of DOPA and 5-hydroxy-tryptophan is catalyzed by the same enzyme, DDC. In vertebrates this enzyme is called amino acid decarboxylase (AADC). Only vertebrates further metabolize dopamine to norepinephrine and epinephrine. TH, tryosine hydroxylase DDC, DOPA decarboxylase DBH, dopamine b-hydroxylase PNMT, phenylethanolamine N-methyltransferase. Tryp-OH tryptophan hydroxylase.

A common way to benefit from the ability to combine different molecular orbital methods in ONIOM is to combine a DFT or ab-initio description of the reactive region with a semi-empirical treatment of the immediate protein environment, including up to 1000 atoms. Due to the requirement for reliable semi-empirical parameters, as discussed in Section 2.2.1, this approach has primarily been used for non-metal or Zn-enzymes. Examples include human stromelysin-1 [83], carboxypeptidase [84], ribonucleotide reductase (substrate reaction) [85], farnesyl transferase [86] and cytosine deaminase [87], Combining two ab-initio methods of different accuracy is not common in biocatalysis applications, and one example from is an ONIOM (MP2 HF) study of catechol O-methyltransferase [88],... 

With the characterized mechanism, the next key question is the origin of its catalytic power. A prerequisite for this investigation is to reliably compute free energy barriers for both enzyme and solution reactions. By employing on-the-fly Born-Oppenheimer molecular dynamics simulations with the ab initio QM/MM approach and the umbrella sampling method, we have determined free energy profiles for the methyl-transfer reaction catalyzed by the histone lysine methyltransferase SET7/9... 

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