Sulfatase regulation and antibiotic synthesis in Cephalosporium acremonium

An early blocked sulfur amino acid auxotroph, Cephalosporium acremonium mutant 274-1 (which could be satisfied by methionine or cysteine), utilized organic sulfur compounds for cephalosporin C production in the following order of decreasing effectiveness; methionine > cystathionine > cysteine, despite the fact that cysteine is considered to be the immediate precursor of the antibiotic. When a genetic block was added to mutant 274-1 in the transsulfuration pathway from cysteine to methionine, the double mutant 11-8 (which grows on methonine but not cysteine) failed to produce cephalosporin C from cysteine even though enough methionine was added to support normal growth. Addition of the non-sulfur analogue, norleucine, resulted in antibiotic production from cysteine in the double mutant. These facts support the hypothesis that methionine stimulation of cephalosporin C production is due to a role of methionine other than that of sulfur donation.The probable pathway of sulfur metabolism in Cephalosporium acremonium is shown in Fig. 1. Although the immediate donor of sulfur to cephalosporin C is cysteine, this amino acid exerts little to no stimulatory effect on antibiotic production in a sulfate-containing medium. Methionine, on the other hand, is an excellent stimulator (for review, see reference 2). We suspect that the effect of methionine is a regulatory one. We have recently shown (6) that methionine has an obligatory role in stimulation of cephalosporin C production from sulfate even though the shortest path from sulfate to cephalosporin C (Fig. 1) does not include methionine. Using a methionine auxotroph (S-1), presumably blocked in transsulfuration between cystationine and homocysteine, we found that limitation of methionine in the presence of excess sulfate inhibited antibiotic production but not growth; i.e., the mutant produced no cephalosporin C from sulfate whereas its parent did produce the antibiotic. Although our calculations indicated that the small amount of methionine added for growth of mutant S-1 would not be expected to shut off sulfate assimilation, we felt that the known depressive effect of methionine on sulfate uptake (1) constituted an unnec-' Contribution no. 2565 from the

Section: Discussionmentioning

confidence: 99%

Production of Cephalosporin C by Single and Double Sulfur Auxotrophic Mutants of Cephalosporium acremonium

Drew

Demain

1975

“…73, p. 37,1971.) The derivation of the M8650 mutant series of C. acremonium (reclassified as Acremonium chrysogenum by W. Gams [4]) has been previously described (2,6). Alternatively, mutants were screened for the inability to synthesize cephalosporin C by extracting a standard volume of agar beneath each colony and assaying the extract with Alcaligenes faecalis ATCC 8750 in the presence of penicillinase.…”

mentioning

confidence: 99%

“…Alternatively, these antibiotics were separated by paper chromatography using a propanolpyridine-acetic acid-acetonitrile-water (45:30:9:40:36) mixture or a methanol-n-propanol-water (6:2:1) mix- (9). The difference between untreated and penicillinase-treated samples measured the amount of penicillin N. In addition, penicillin N was determined biologically by a plate assay previously described (2) and by placing the dried paper chromatograms on plates seeded with Salmonella gallinarum and plates seeded with S. gallinarum plus penicillinase. Penicillin N zones were compared with standards.…”

mentioning

confidence: 99%

Synthesis of Deactoxycephalosporin C by a Mutant of Cephalosporium acremonium

Queener

Capone

Radue

et al. 1974

A number of Cephalosporium acremonium mutants blocked in the synthesis of cephalosporin C were investigated for accumulation of other ,3-lactam compounds. The non-cephalosporin C producers were isolated after exposing the superior cephalosporin C-producing strain M8650-4 to ultraviolet light (268 nm). One of the blocked mutants, MH63, accumulated deacetylcephalosporin C (0.4 mg/ml), deacetoxycephalosporin C (1.5 mg/ml), and penicillin N (2.7 mg/ml). In contrast, the parent of MH63 produced high levels of cephalosporin C as well as deacetylcephalosporin C (2.2 mg/nal) and penicillin N (1.0 mg/ml), but only traces of deacetoxycephalosporin C (about 0.1 mg/ml). Deacetoxycephalosporin C was isolated from the mutant strain and identified by ultraviolet light, nuclear magnetic resonance, bioactivity spectrum, and co-migration with authentic standard in three chromatography systems.

“…Mutant M8650-sp-1 was atypical when compared with other strains of the M8650 series (6). Strains previously assigned to this series exhibit increased potential for synthesis of antibiotic in methionine-medium as well as in sulfate-medium (Table 2).…”

Section: Methodsmentioning

confidence: 96%

“…Methionine-medium contained 0.5% DL-methionine in place of calcium sulfate, and the content of calcium carbonate in this medium was adjusted to 0.6%. Cells also were grown in a synthetic medium (6) and were used to study the uptake of sulfate. Cultures were grown either in 500-ml Erlenmeyer flasks containing 60 Dennen and Carver (6).…”

Section: Methodsmentioning

confidence: 99%

Synthesis of Cephalosporin C from Sulfate by Mutants of Cephalosporium acremonium

Niss

Nash

1973

The innate ability of Cephalosporium acremonium to use methionine preferentially over sulfate for synthesis of cephalosporin C can be influenced through mutation. Mutants of C. acremonium with altered capacity to utilize sulfate for synthesis of antibiotic were isolated and partially characterized with respect to the uptake of sulfate and the regulation of arylsulfatase.