Anthony Joseph Tietz

The formation of daptomycin by supplying decanoic acid to Streptomyces roseosporus cultures producing the antibiotic complex A21978C

Antibiotic substance A21978C is a complex of compounds having a common cyclic polypeptide nucleus and different fatty acid sidechains. Daptomycin is a semi-synthetic antimicrobial substance derived from the A21978C complex by a very elaborate chemical process. To obtain the daptomycin, the A21978C complex was first isolated from culture filtrates of Streptomyces roseosporus by several resin procedures. The resulting material was then ‘blocked’ and added to an Actinoplanes culture for deacylation. The protected A21978C nucleus was subsequently isolated by the same procedure as the parent complex and reacylated with the desired fatty acid (decanoic acid). The acylated compound was deblocked to yield daptomycin. This report describes the experimentation undertaken to establish a strategy to supply a very toxic precursor to cultures of S. roseosporus and, thereby, produce daptomycin biosynthetically.

TRANSPOSITION MUTAGENESIS IN STREPTOMYCES FRADIAE : IDENTIFICATION OF A NEUTRAL SITE FOR THE STABLE INSERTION OF DNA BY TRANSPOSON EXCHANGE

We explored transposition in Streptomyces fradiae (Sf) as a means to insert a second copy of the tylF gene to improve tylosin (Ty) production. Transposons Tn5096 and Tn5099 transposed relatively randomly in Sf, and many of the insertions caused no deleterious effects on Ty production yields. Tn5098, a derivative of Tn5096 containing tylF and tylJ genes, recombined into the chromosome into the tyl gene cluster and transposition was not observed. However, following the tagging of a neutral site (NS) by Tn5099 transposition, tylF was effectively inserted into the NS by homologous recombination (transposon exchange). Recombinants obtained by transposon exchange produced higher yields of Ty.

High yielding culture conditions for the biosynthesis of D-amino acid oxidase byTrigonopsis variabilis

SummaryA process was developed for the production ofTrigonopsis variabilis and D-amino acid oxidase. Yields for the yeast were in excess of 220 g/l wet weight and 62 g/l dry weight. Using cephalosporin C as a substrate the enzyme concentration was 7000 units per liter.

Defined media strategies for the biosynthesis of Cephalosporin C

SummaryA stirred reactor process is described for the production of Cephalosporin C in high yields. Using a chemically defined medium, fermentation control strategies were developed to optimize the production of the antibiotic. The highest yields were obtained with carbon as the limiting substrate during both the rapid growth and synthetic phases of the process; the relative merits of glucose and lipid as carbon sources were examined.

The synthesis of A21978C analogs byStreptomyces roseosporus cultivated under carbon limitation and fed fatty acids

SummaryBy employing carbon limited fed batch techniques, fatty acids of length C8 to C12 were successfully employed as precursors for biosynthesis of A21978C antibiotics. The efficiency of incorporation was related to the length of the lipid precursor, with C10-C12 fatty acids being the preferred substrates.

Characterization of the process for the biosynthesis of the actaplanin complex by Actinoplanes missouriensis

Abstract The production of the antibiotic actaplanin by Actinoplanes missouriensis was characterized in a high yielding process. The data generated, indicate that the antibiotic is synthesized during a period of declining cell mass as estimated by dry weight and viability. Further study demonstrated that the producing culture binds actaplanin and is inhibited by the antibiotic in the soluble form.