S-Adenosyl- L-Methionine And S-Adenosyl- L-Homocysteine: A Nuclear Magnetic Resonance Study And Purification Of S-Adenosyl- L-Methionine: L-Homocysteine S-Methyltransferase From Saccharomyces Cerevisiae

• Main Author: Stolowitz, Mark Lewis
• Format: Others
• Published: Scholarly Commons 1981
• Subjects: Biochemistry; Pure sciences; Physical Sciences and Mathematics
• Online Access: https://scholarlycommons.pacific.edu/uop_etds/3158; https://scholarlycommons.pacific.edu/cgi/viewcontent.cgi?article=4165&context=uop_etds

An analysis of the 360 MHz ('1)H NMR spectra of the title compounds in ('2)H(,2O) is presented. The ('3)J values for the ribose vicinal protons of S-adenosyl-L-methionine are consistent with a predominantly C(,3')-exo conformation and with one highly favored gauche-anti conformation about the C(,4')-C(,5') bond. The corresponding ('3)J values for S-adenosyl-L-homocysteine imply a similar C(,3')-exo ribose ring conformation, but the orientation about the C(,4')-C(,5') bond is distributed between two gauche-anti rotamers. The methionine side chain of S-adenosyl-L-methionine has approximately equal populations of rotational isomers about the C(,(alpha))-C(,(beta)) and C(,(beta))-C(,(gamma)) bonds, whereas the side chain of S-adenosyl-L-homocysteine exhibits a conformational preference for the gauche-anti conformations about the C(,(alpha))-C(,(beta)) bond. ('1)H and ('13)C NMR spectra of commercially available samples of (-)S-adenosyl-L-methionine consistently available samples of (-)S-adenosyl-L-methionine consistently reveal the presence of a small amount of the (+) sulfonium diastereomer. This assignment was confirmed by the synthesis of both the ('1)H and ('13)C methyl derivatives of S-adenosyl-L-homocysteine. Arguments are presented to explain the failure of previous workers to detect (+)S-adenosyl-L-methionine in biological preparations. The possible biological significance of this finding is discussed with reference to the enzyme S-adenosyl-L-methionine: L-homocysteine S-methyltransferase which employs both (-)S-adenosyl-L-methionine and (+)S-adenosyl-L-methionine as methyl donors. An improved purification of S-Adenosyl-L-methionine: L-homocysteine S-methyltransferse (EC. 2.1.1.10) from Saccharomyces cerevisiae is reported. The enzyme was purified approximately 1500-fold by toluene extraction, ammonium sulfate precipitation, Sephadex G25 gel exclusion chromatography, DEAE Sephadex ion-exchange chromatography and affinity chromatography on L-methionine AH-Sepharose 4B. The procedure affords a 12-fold increase in yield and a 3-fold increase in purification over the previous fractionation scheme. It is hoped that the availability of a highly purified preparation of the enzyme will provide the basis for an investigation of enzymatic transmethylation in which the fate of the sulfonium diastereomers of S-adenosyl-L-methionine is directly observable by NMR spectroscopy.