Aureothricin

L-Methionine and DL-ethionine decreased production of thiolutin and aureothricin in Streptomyces kasugaensis. In the presence of L-methionine the culture also produced 3-methylthioacrylic acid, 3-methylthiopropionic acid and 3,6-bis-(2-methylthioethyl)-2,5-dioxopiperazine. Production of the metabolites depended on the concentration of L-methionine in the medium.

Holomycin is a member of the dithiolopyrrolone class of secondary metabolites. It contains a cromophore formed by two heterocycles, one of them carrying a disulfide bridge. The holomycin chromophore is also present in thioluthin, aureothricin, the xenorhabdins and the complex thiomarinols. Streptomyces clavuligerus ATCC 27064 contains the holomycin gene cluster (hlm). This antibiotic is formed by a cysteine-activating non-ribosomal peptide synthetase followed by four reduction steps by a set of four different flavoproteins; the intermediate is cyclized by a thiol oxidase and modified by acylation. Holomycin is a broad spectrum antibiotic, reported as antitumoral, acting in vivo on RNA synthesis. It is modified intracellularly by the producer strains by methylation and formation of heterodimers as a way of self-protection. Holomycin might be a lead molecule for the production of new hybrid compounds with higher activity and lower toxicity.

The spontaneous mutant 18a derived from Streptomyces kasugaensis MB273 exhibited pleiotropic effect such as loss of aerial mycelium formation, aureothricin (AT) production, and of citrullin biosynthesis, as well as changes in plasmid; the mutant required cystine for production of aureothricin. An improved method of protoplast regeneration was applied to S. kasugaensis MB 273-18a and a regeneration efficiency of 90% or more was obtained. Sixty to ninety percent of the colonies regenerated from the 18a protoplasts exhibited reversion of the pleiotropic mutation in 18a. Moreover, of 13 regenerated strains which showed these drastic phenotypic variations, it was found that their plasmid types varied. These types could be divided into two groups; the RI type (5 strains) which contained a large amount of pSK2, a small amount of pSK3 and no pSK1, and the RII type (8 strains) in which no closed-circular DNA was detected. From these results, the following conclusions were obtained. First, plasmid curing in RII type strains and also the variation of plasmid copy in the RI type strains occurred as the result of protoplast regeneration. Second, the structural genes for biosynthesis of AT probably exist on chromosome. Third, regeneration of 18a protoplasts causes the reversion of pleiotropic mutation with high frequency. A working hypothesis was proposed to explain these complex phenomena.