This invention relates to recombinant DNA technology. In particular the invention pertains to the cloning of the SAM operon genes from Streptomyces fradiae and the use of said genes and their encoded proteins to produce S-adenosylmethionine (SAM) in a recombinant host.
S-adenosylmethionine is a product of natural origin found in all living organisms. SAM is a product of considerable importance for its role in biological reactions such as transmethylations. While the enzymes that catalyze these reactions are varied in their substrate specificity they are practically universal in their requirement of S-adenosylmethionine as the ultimate methyl group donor. Some methyl transfer reactions are important in the synthesis of certain antibiotics, such as tylosin.
Tylosin is a macrolide antibiotic composed of a 16-membered branched lactone, tylactone, and residues of three attached sugars, mycaminose, mycarose, and mycinose. Tylosin is produced commercially by Streptomyces fradiae (ATCC 19609; NRRL 2702) and is used as an animal growth promotant and veterinary antibiotic. The multi-step biosynthesis of tylosin has been studied both physiologically and genetically (See generally, R. H. Baltz and E. T. Seno, "Genetics of Streptomyces fradiae and tylosin biosynthesis. Ann. Rev. Microbiol. 42, 547-74 (1988)). At least 13 biosynthetic genes and 2 regulatory genes are necessary for normal production of tylosin. Tylosin synthesis requires multiple methylation reactions, the last two of which are rate-limiting. In the last step a specific methyltransferase catalyzes the transfer of a methyl group from SAM to the tylosin precursor molecule, macrocin. Thus, the availability of SAM as the methyl group donor is essential in the synthesis of tylosin.
S-adenosylmethionine is produced when an adenosyl group is transferred from ATP to methionine. SAM is synthesized in the cell by the action of three enzymes encoded by the SAM operon--SAM synthetase, methyl transferase (MT), and methylene tetrahydrofolate reductase (MTHR).