Methyltransferase methyl transfer

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... 

In cells that synthesize epinephrine, the final step in the pathway is catalyzed by the enzyme phenylethanolamine /V-methyltransferase. This enzyme is found in a small group of neurons in the brainstem that use epinephrine as their neurotransmitter and in the adrenal medullary cells, for which epinephrine is the primary hormone secreted. Phenylethanolamine N-methyltransferase (PNMT) transfers a methyl group from S-adenosylmethionine to the nitrogen of norepinephrine, forming a secondary amine [5]. The coding sequence of bovine PNMT is contained in a... 

Histone-lysine methyltransferases are chromatin-bound enzymes that catalyses the addition of methyl groups onto lysine or arginine residues of chromatin-bound H3 and H4 [151]. The methyl group is transferred enzymatically to the histone with S-adenosyl methionine as the methyl donor. Histone methylases have been isolated from HeLa S-3 cells [182], chick embryo nuclei [183], and rat brain chromatin [184]. The histone methyltransferases methylated H3 and H4 in nucleosomes [184]. Histone-lysine methyltransferase is a chromatin-bound enzyme [129,151]. Initial characterization of the Tetrahymena macronuclear H3 methyltransferase suggests that the enzyme has a molecular mass of 400 kDa. The enzyme preferred free histones rather than nucleosomes as substrate [138]. More recent studies have now... 

It should be noted, however, that these enzymes catalyzed single methylation steps, and did not accept partially methylated substrates for further O-methylation. The common occurrence of partially methylated flavonoids (16,17), such as those of C. americanum (Figure 1), raised the question as to whether multiple methyl transfers were catalyzed by one or several position-specific O-methyltransferases ... 

Wagner et al. (22) isolated Pseudomonas MS which grew on trimethylsul-phonium salts as a sole source of carbon and energy, with the evolution of DMS. A partially purified enzyme preparation catalyzed the transfer of a methyl group from trimethylsulfonium chloride to tetrahydrofolate. Neither S-aaenosyl-methionine nor DMSP functioned in the methyl transfer reaction. The methyltransferase was devoid of vitamin B12, had a molecular weight of about 100,(JOO with a pH optimum of 7.8. 

Protein lysine methyltransferases (PKMTs) are a family of enzymes that transfer the activated methyl group from S-adenosyl-L-methionine (SAM) to specific lysine residues on various substrates. The PKMTs have been causally linked to various human diseases including cancer [140], Huntington s disease [141], and growth defects [142, 143]. The substrates of the PKMTs are typically histones [144-146], but there are several methyltransferases methylate non-histone substrates, such as the tumor suppressor p53 [147, 148], the estrogen receptor ERa [149], and the ATPase Reptin [150]. Given the importance of these enzymes in normal and... 

Hegazi, M. F. Borchardt, R. T. Schowen, R. L. a-Deuterium and carbon-13 isotope effects for methyl transfer catalyzed by catechol 0-methyltransferase. Sj 2-like transition state, J. Am. Chem. Soc. 1979,101,4359-4365. 

Ounaroon A, Decker G, Schmidt J, Lottspeich F, Kutchan TM. (R,S)-reticuline 7-0-methyltransferase and (R,S)- norcoclaurine 6-O-methyltransferase of Papaver somniferum - cDNA cloning and characterization of methyl transfer enzymes of alkaloid biosynthesis in opium poppy. Plant J. 2003 36 808-819. 

Ferguson DJ Jr, Gorlatova N, Grahame DA, Krzycki JA. Reconstitution of dimethylamine coenzyme M methyl transfer with a discrete corrinoid protein and two methyltransferases purified from Methanosarcina barker . J. Biol. Chem. 2000 275 29053-29060. Hao B, Gong W, Ferguson TK, James CM, Krzycki JA, Chan MK. A new UAG-encoded residue in the structure of a methanogen methyltransferase. Science 2002 296 1462-1466. 

Burke SA, Krzycki JA. Reconstitution of Monomethylamine Coenzyme M methyl transfer with a corrinoid protein and two methyltransferases purified from Methanosarcina barkeri. J. Biol. Chem. 1997 272 16570-16577. 

Cobamides or corrinoids are involved in methyl-transfer reactions in the methanogenic pathways, especially from methyl substrates [143-156]. At least one cobamide is involved in methanogenesis from H2-CO2 by M. thermoautotrophicum, where it is found in the methyl-H4MPT CoM methyltransferase [157]. The majority are either associated with membranes or bound to soluble proteins [143,158]. Methanogens are inhibited by corrinoid antagonists, suggesting an important metabolic role [159]. 

Two models have been proposed for this methyl-transfer reaction, (i) Recent evidence from Kengen et al. suggests that the methyl group of CH3-H4MPT is first transferred to a corrinoid protein called methyltransferase a (MTa) yielding protein-bound CH3-B12 HBI (Reaction 19) which acts as the methyl-donor to CoM in a reaction catalyzed by methyltransferase MT2, a non-corrinoid protein (Reaction 20) [157] ... 

The oxygen-insensitive MT2 protein was previously purified by Taylor and Wolfe as methylcobalamin CoM methyltransferase from Methanobactehum bryantii [194]. Reactions (19) and (20) are analogous to steps in methyl transfer from methanol to CoM. 

M. barkeri extracts methanogenesis from this labelled protein is at least 100 times faster than the non-enzymatic reaction between CoM and methylcobalamin they postulated that this protein acts as a methyltransferase in methanogenesis from methanol. Finally, Van der Meijden et al. [152-155] resolved the M. barkeri methyl transfer from methanol to CoM into two steps involving methyltransferase 1 and 2, or MTi and MT2. The oxygen-sensitive corrinoid protein MTj in its most reduced state (Co" ) accepts the methyl group from methanol [153,154] ... 

The following data indicate that CH3-H4MPT H-S-CoM methyltransferase is the site of primary Na translocation (see Figs. 6 and 12) (i) the enzyme has been partially purified from Methanosarcina barkeri and Methanobacterium thermoautotrophicum and found to be tightly membrane bound [69b] (ii) inverted vesicles of the methanogenic strain G61 catalyzed methyl transfer from CH3-H4MPT to H-S-CoM. This reaction was stimulated by Na ions and was coupled with the accumulation of Na" into the vesicles. Na uptake was inhibited by Na ionophores rather than by protonophores indicating primary Na translocation [168]. 

TaUant T. C., Paul L., and Krzycki J. A. (2001) The MtsA subunit of the methylthiol coenzyme M methyltransferase of Methanosarcina barkeri catalyses both half-reactions of corrinoid-dependent dimethylsuUide coenzyme M methyl transfer. J. Biol. Chem. 276, 4485—4493. 

An extremely interesting application of KIEs and theory has been elucidating the effect of enzyme catalysis on the structure of the SN2 transition state. Schowen and coworkers29 30 measured the secondary a-deuterium and a-carbon 12C/13C KIEs for the enzyme-catalyzed SN2 methyl transfer reaction between G-adenosylmethionine (Fig. 18) and 3,4-dihydroxyacetophenone in the presence of the rat-liver enzyme catechol O-methyltransferase (COMT) at 37°C (Fig. 19) and for the closely related, uncatalyzed Sn2 reaction between methoxide ion and G-methyldibenzothiophenium ion in methanol at 25°C (Fig. 20). The near maximum a-carbon KIEs of 1.09 + 0.02 for the enzyme-catalyzed SN2 reaction and 1.08 + 0.01 for the uncatalyzed Sn2 reaction were taken as evidence that both transition states were symmetric. However,... 

As observed in the section discussing BtrN and DesII, radical SAM enzymes have already been shown to play a critical role in the synthesis of certain elements of antibiotic compounds. Moreover, we observed in the MiaB section that a radical SAM enzyme is responsible for catalyzing a methylation reaction in the synthesis of ms i A. Methyl transfer reactions in biology commonly utilize DNA methylase enzymes or SAM-dependent methyltransferases the latter system has been touched upon in the multiple SAM-dependent methyl transfer reactions involved in yW synthesis. However, a unique methylation reaction has recently been proposed that invokes the use of both SAM and methylcobalamin. 

The methyltransferase from Haemophilus haemolyticus methylates its recognition sequence, 5 -GCGC-3 in dsDNA to yield 5 -G-m CGC-3. SAM donates the methyl group to become S-adenosylhomoserine. The methyl transfer via formation of the methylated intermediate is initiated by the nucleophilic thiolate attack of Cys81, assisted by the general acid Glull9. Nearly all base specific interactions are made in the major... 

Methyl transfer reactions play a significant part in the modifications of aromatic polyketides, both of the polyketide core [61,62] as well as of several of the sugar moieties [44,53]. In Streptomyces, more than 20 amino acid sequences have been found that may represent enzymes involved in methyl transfer reactions in the biosynthesis of aromatic polyketides [149]. One of these enzymes, the S-adenosyl-L-methionine-dependent DnrK, is involved in the methylation of the C-4 hydroxyl group in daunorubicin/doxorubicin biosynthesis (Scheme 10, step 12). The subunit of the homo-dimeric enzyme displays a fold typical for small-molecule methyltransferases. The structure of the ternary complex with bound products S-adenosyl-L-homocysteine and 4-methoxy-8-rhodomycin provided insights into the structural basis of substrate recognition and catalysis [149]. The position and orientation of the substrates suggest an Sn2 mechanism for methyl transfer, and mutagenesis experiments show that there is no catalytic base in the vicinity of the substrate. Rate enhancement is thus most likely due to orientational and proximity effects [149]. 

The available information on 5-methylmethionine does not provide convincing indication of a physiological role for this compound. If there is any methylated metabolite the formation of which occurs only through utilization of 5-methylmethionine as methyl donor, that fact has yet to be discovered. For the moment, plant methyltransferases appear to conform to the generalization that for an enzymatic methyl transfer to a compound other than homocysteine, AdoMet is the methyl donor (Mudd and Cantoni, 1%4 Canton , 1977). Although 5-methylmethionine has been tested as a methyl donor with only a minority of the reported plant AdoMet-dependent methyltransferases, in those cases in which it has been tried it has been inactive (Mudd, l%0b Mann et al., 1963). 

Zhao, S. Y Ragsdale, S. W. A conformational change in the methyltransferase from Clostridium thermoaceticum facilitates the methyl transfer from (6S)-methyltetrahydro-folate to the corrinoid iron-sulfur protein in the acetyl-CoA pathway. Biochemistry, 1996, 35(7), 2476-2481. 

C-methylation reaction is the methylation of the C-5 position of cytosine in DNA. In this case, the carbon C-5 of cytosine cannot directly act as a nucleophile. The electron withdrawal by N-3 and the carbonyl, however, makes the C-5—C-6 double bond electron deficient and prone to attack by nucleophiles in a reaction that is similar to a Michael reaction. In DNA methyltransferases (DNMTs), this nucleophile is the thio-late from a Cys residue. The addition product is nucleophilic and reacts with SAM via an Si,j2-like mechanism to capture the methyl group. The resulting intermediate then eliminates the Cys of DNMT to give the methylated cytosine product (Figure 1.9). The methylation of C-5 of cytosine is an example of converting an electron-deficient methyl acceptor to a nucleophile for the methyl-transfer reaction by addition of an active site Cys thiolate. 

In the proposed biosynthesis pathway of wybutosine, the acp-transfer step is catalyzed by Tyw2, which has similarity to methyltransferases that catalyze nucleophilic methyl-transfer reactions (Figure 1.11). 

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