Methionine activating enzyme

Norepinephrine is methylated in the presence of a specific enzyme (S-adenosylmethionine transferase) and a cofactor (S-adenosylmethionine) to form epinephrine. This reaction is analogous to the methyla-tion of guanidinoacetic acid, which occurs in liver. Two distinct enzyme reactions are involved methionine is converted to S-adenosylmethionine in the presence of a methionine-activating enzyme and ATP, in which reaction all the phosphates of ATP are lost, the terminal phosphorus of ATP is liberated as inorganic phosphate and the two internal phosphoryl groups yield pyrophosphate and (2) norepinephrine is then methylated in the presence of a specific enzyme S-adenosylmethionine transferase to form epinephrine. 

Because of its role in activating methionine for transmethylation, this enzyme was initially called the methionine-activating enzyme. With the later discovery of enzymes which activate the carboxyl group of methionine and other amino acids for protein synthesis, this term became somewhat confusing. In this chapter, the trivial name methionine adenosyltransferase or adenosyhransferase, will be used in accord with the recommendation of the Commission on Enzymes (Enzyme Nomenclature, American Elsevier, New York, 1965). The enzyme has also been called S-adenosylmethionine synthetase. 

Isolation of alkaline phosphatase from Escherichia coli in which 85% of the proline residues were replaced by 3,4-dehydro-proline affected the heat lability and ultraviolet spectrum of the protein but the important criteria of catalytic function such as the and were unaltered (12). Massive replacement of methionine by selenomethionine in the 0-galactosidase of E. coli also failed to influence the catalytic activity. Canavanine facilely replaced arginine in the alkaline phosphatase of this bacterium at least 13 and perhaps 20 to 22 arginyl residues were substituted. This replacement by canavanine caused subunit accumulation since the altered subunits did not dimerize to yield the active enzyme (21). Nevertheless, these workers stated "There was also formed, however, a significant amount of enzymatically active protein in which most arginine residues had been replaced by canavanine." An earlier study in which either 7-azatryptophan or tryptazan replaced tryptophan resulted in active protein comparable to the native enzyme (14). 

The sulfur atom of methionine residues may be modified by formation of sulfonium salts or by oxidation to sulfoxides or the sulfone. The cyanosulfonium salt is not particularly useful for chemical modification studies because of the tendency for cyclization and chain cleavage (129). This fact, of course, makes it very useful in sequence work. Normally, the methionine residues of RNase can only be modified after denaturation of the protein, i.e., in acid pH, urea, detergents, etc. On treatment with iodoacetate or hydrogen peroxide, derivatives with more than one sulfonium or sulfoxide group did not form active enzymes on removal of the denaturing agent (130) [see, however, Jori et al. (131)]. There was an indication of some active monosubstituted derivatives (130, 132). 

Many crystal structures of other potassium-activated enzymes have been reported. They generally require a divalent cation and are activated by a monovalent cation that also binds. Such enzymes include amylase, which has a unique Ca Na a trinuclear center, 5 -adenosyImethionine synthetase, E. coli methionine aminopeptidase in which... 

Type 2 copper centers are not uniform in ligand or ligand stereochemistries. One common feature is, however, that in the active enzyme, one coordination site is always free to bind oxygen. The most common ligand in type 2 copper centers is histidine. Tyrosine (often modified), methionine, and cysteine occur as well. There are three histidines and a modified tyrosine in amine oxidase and lysyl oxidase [28]. In diamine oxidase, two of the histidine residues have probably been replaced by cysteines [29]. In galactose oxidase, the copper ion is coordinated by two tyrosines, two histidines and an acetate ion [30]. Dopamine-/J-hydroxylase contains two differently coordinated copper ions per functional unit. One is coordinated by three histidines and a methionine and the other by two histidines and another, yet unknown, ligand [ 31 ]. Last but not least, the type 2 copper ion in Cu,Zn-superoxide dismutase is coordinated by four histidine residues, one of which connects the copper ion to the zinc ion, the second metal ion in the active site of the enzyme [32,33] (Fig. 6). 

The enzyme is a homodimeric protein of A/r 170,000 and contains no known organic or metal ion cofactors. The enzyme is readily inactivated by oxygen and interconverts between active and inactive forms in vivo (173, 174). The activation process occurs under conditions of anaerobiosis and is catalyzed by an Fe(ll)-dependent activating enzyme (Mr 30,000) (775). Elegant studies on the in vitro activation of PFL by Knappe and co-workers (176, 177) have revealed that a complex activation cocktail is required, which includes the activating enzyme, pyruvate, or oxamate as allosteric effectors, S-adenosylmethionine (SAM), and flavodoxin (775) or photoreduced 5-deazariboflavin (178). A possible role for a B12 derivative in the activation or catalytic reaction for PFL is not likely in light of the observation that E. coli 113-3, a methionine/B auxotroph, pos-... 

Figure 15 Cleavage of S-adenosyl-L-methionine by enzymes within the radicai SAM superfamiiy. The reductive cleavage reaction generates a 5 -deoxyadenosyl 5 -radical and L-methionine, a spectator in the reaction. The physiological electron donor is the flavodoxin/flavodoxin reductase reducing system, with electrons deriving ultimately from NADPH. Artificial electron donors such as sodium dithionite and 5-deazaflavin plus light can also effect reduction in in vitro activity determinations.

In E. coli, ThiH catalyzes the formation of the glycine imine 23 from tyrosine (26). ThiH is an oxygen-sensitive radical 5-adenosyl-L-methionine (SAM) enzyme. Its activity has been reconstituted and the mechanism outlined in Figure 8 has been proposed. It is unclear why E. coli adopts such a complex route to the glycine imine when oxidation of glycine using nicotinamide adenine dinucleotide (NAD) would accomplish the same transformation. 

As the name implies, renal clearance of abnormal levels of homocystine in the plasma causes excessive excretion of the amino acid in the urine. In cystathionine P-synthase deficiency, plasma methionine concentrations are elevated as well -this serves as a point of distinction from the remethylation defects. At present, it appears that the pyridoxal phosphate response may be explained by the fact that this vitamin increases the steady-state concentration of the active enzymes by decreasing the rate of apoenzyme degradation and possibly by increasing the rate of holoenzyme formation. The explanation is not entirely satisfactory, however, since in vitro studies have shown detectable levels of enzyme activity in mutant fibroblasts that have no response, while in other mutant lines without detectable enzyme activity, response has occurred. Once again, a distressing lack of correspondence between in vivo observations and in vitro experiments forces investigators to probe the secrets of these diseases more deeply. 

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