Synthesis and characterization of 2-mercaptoethanesulfonic acid albumin.

Autoimmune patients treated with ifosfamide (CAS 3778-73-2) and mesna (2-mercaptoethanesulfonic acid, CAS 3375-50-6) in some cases suffered from severe allergic reactions that were proposed to be due to mesna linked to serum albumin by a disulfide bond. To prove the existence of the hypothetic mesna albumin adduct in vivo it was synthesized: The free thiol group of albumin (molecular mass determined by MALDI spectroscopy: 67009 Da) was converted to S-phenylsulfonyl albumin and reacted with mesna to albumin mesna (molecular mass: 67159 Da). In an alternative synthesis albumin was incubated with mesna at pH 8, 40 degrees C (molecular mass of the adduct: 67166 Da).

Prebiotic syntheses of vitamin coenzymes: I. Cysteamine and 2-mercaptoethanesulfonic acid (coenzyme M).

The reaction of NH3 and SO3(2-) with ethylene sulfide is shown to be a prebiotic synthesis of cysteamine and 2-mercaptoethanesulfonic acid (coenzyme M). A similar reaction with ethylene imine would give cysteamine and taurine. Ethylene oxide would react with NH3 and N(CH3)3 to give the phospholipid components ethanolamine and choline. The prebiotic sources of ethylene sulfide, ethylene imine and ethylene oxide are discussed. Cysteamine itself is not a suitable thioester for metabolic processes because of acyl transfer to the amino group, but this can be prevented by using an amide of cysteamine. The use of cysteamine in coenzyme A may have been due to its prebiotic abundance. The facile prebiotic synthesis of both cysteamine and coenzyme M suggests that they were involved in very early metabolic pathways.

Transport of coenzyme M (2-mercaptoethanesulfonic acid) and methylcoenzyme M [(2-methylthio)ethanesulfonic acid] in Methanococcus voltae: identification of specific and general uptake systems.

A transport system for coenzyme M (2-mercaptoethanesulfonic acid [HS-CoM]) and methylcoenzyme M [(2-(methylthio)ethanesulfonic acid (CH3-S-CoM)] in Methanococcus voltae required energy, showed saturation kinetics, and concentrated both forms of coenzyme M against a concentration gradient. Transport required hydrogen and carbon dioxide for maximal uptake. CH3-S-CoM uptake was inhibited by N-ethylmaleimide and monensin. Both HS-CoM and CH3-S-CoM uptake showed sodium dependence. In wild-type M. voltae, HS-CoM uptake was concentration dependent, with a Vmax of 960 pmol/min per mg of protein and an apparent Km of 61 microM. Uptake of CH3-S-CoM showed a Vmax of 88 pmol/min per mg of protein and a Km of 53 microM. A mutant of M. voltae resistant to the coenzyme M analog 2-bromoethanesulfonic acid (BES) showed no uptake of CH3-S-CoM but accumulated HS-CoM at the wild-type rate. While the higher-affinity uptake system was specific for HS-CoM, the lower-affinity system mediated uptake of HS-CoM, CH3-S-CoM, and BES. Analysis of the intracellular coenzyme M pools in metabolizing cells showed an intracellular HS-CoM concentration of 14.8 mM and CH3-S-CoM concentration of 0.21 mM.

Substrate analogues as mechanistic probes of methyl-S-coenzyme M reductase.

Methyl-S-coenzyme M reductase catalyzes the ultimate methane-yielding reaction in methanogenic bacteria, the reductive cleavage of the terminal carbon-sulfur bond of 2-(methylthio)ethanesulfonic acid. This protein has previously been shown to contain 2 equiv of a tightly bound nickel corphinoid cofactor, denoted cofactor F430, that may play a role in catalysis. Prior to this study, only one substrate analogue, ethyl-S-coenzyme M, had been demonstrated to be processed to a product by anaerobic cell extracts from Methanobacterium thermoautotrophicum strain delta H. In this investigation, we have synthesized three additional substrate analogues that serve as substrates as well as five previously unknown inhibitors. Steady-state kinetic techniques were developed in order to assess relative rates of processing for these substrates and inhibitors by use of anaerobic cell extracts from M. thermoautotrophicum. With this assay system, a KM of 0.1 mM and a kcat of 17 min-1 were determined for methyl-S-coenzyme M as substrate. Methyl-seleno-coenzyme M was converted to methane with a kcat threefold higher than that of methyl-S-coenzyme M, but kcat/KM was unchanged. The carbon-oxygen bond of 2-methoxyethanesulfonic acid was not cleaved to yield methane, but this analogue acted as an inhibitor with a K1 of 8.3 mM. Methyl reductase catalyzed reductive cleavage of difluoromethyl-S-coenzyme M to yield difluoromethane as the sole product, but trifluoromethyl-S-coenzyme M and trifluoromethyl-seleno-coenzyme M were inhibitors and not substrates.(ABSTRACT TRUNCATED AT 250 WORDS)

Preparation of coenzyme M analogues and their activity in the methyl coenzyme M reductase system of Methanobacterium thermoautotrophicum.

A number of 2-(methylthio)ethanesulfonate (methyl-coenzyme M) analogues were synthesized and investigated as substrates for methyl-coenzyme M reductase, an enzyme system found in extracts of Methanobacterterium thermoautotrophicum. Replacement of the methyl moiety by an ethyl group yielded an analogue which served as a precursor for ethane formation. Propyl-coenzyme M, however, was not converted to propane. Analogues which contained additional methylene carbons such as 3-(methylthio)propanesulfonate or 4-(methylthio)butanesulfonate or analogues modified at the sulfide or sulfonate position, N-methyltaurine and 2-(methylthio)ethanol, were inactive. These analogues, in addition to a number of commercially available compounds, also were tested for their ability to inhibit the reduction of methyl-coenzyme M to methane. Bromoethanesulfonate and chloroethanesulfonate proved to be potent inhibitors of the reductase, resulting in 50% inhibition at 7.9 X 10(6) M and 7.5 X 10(5) M. Analogues to coenzyme M which contained modifications to other regions were evaluated also and found to be weak inhibitors of methane biosynthesis.