Showing papers by "Arnold L. Demain published in 1997"
TL;DR: Production of the antibacterial polypeptide microcin B17 by Escherichia coli ZK650 was inhibited by simulated microgravity, and the site of MccB17 accumulation was found to be different, depending on whether the organism was grown in shaking flasks or in rotating bioreactors designed to establish a simulated micro gravity environment.
Abstract: Production of the antibacterial polypeptide microcin B17 (MccB17) by Escherichia coli ZK650 was inhibited by simulated microgravity. The site of MccB17 accumulation was found to be different, depending on whether the organism was grown in shaking flasks or in rotating bioreactors designed to establish a simulated microgravity environment. In flasks, the accumulation was cellular, but in the reactors, virtually all the microcin was found in the medium. The change from a cellular site to an extracellular one was apparently not a function of gravity, since extracellular production occurred in these bioreactors, irrespective of whether they were operated in the simulated microgravity or normal gravity mode. More probably, excretion is due to the much lower degree of shear stress in the bioreactors. Addition of even a single glass bead to the 50-ml medium volume in the bioreactor created enough shear to change the site of MccB17 accumulation from the medium to the cells.
72 citations
TL;DR: In the new chemically defined medium, which is buffered with 200 mM 2-(N-morpholino)ethanesulfonic acid to prevent decline of pH during fermentation, an ammonium sulfate concentration of 40’mM was optimal for biosynthesis of rapamycin.
Abstract: Six non-amino acid nitrogen compounds were examined as nitrogen source for growth of Streptomyces hygroscopicus and biosynthesis of rapamycin Of the nitrogen sources studied, ammonium sulfate was the best with respect to formation of rapamycin, and supported cell growth comparable to the organic nitrogen sources used in the control chemically defined medium, ie, aspartate, arginine plus histidine In the new chemically defined medium, which is buffered with 200 mM 2-(N-morpholino)ethanesulfonic acid to prevent decline of pH during fermentation, an ammonium sulfate concentration of 40 mM was optimal for biosynthesis of rapamycin Rapamycin production increased by more than 30% on both volumetric and specific bases as compared to the previous medium containing the three amino acids as nitrogen source
66 citations
TL;DR: Examination of gramicidin S production by Bacillus brevis strain Nagano in NASA High Aspect Rotating Vessels found it to be unaffected by simulated microgravity, suggesting the negative effect of glycerol on specific GS formation is dependent on shear and/or vessel geometry, not gravity.
Abstract: In a continuing study of microbial secondary metabolism in simulated microgravity, we have examined gramicidin S (GS) production by Bacillus brevis strain Nagano in NASA High Aspect Rotating Vessels (HARVs), which are designed to simulate some aspects of microgravity. Growth and GS production were found to occur under simulated microgravity. When performance under simulated microgravity was compared with that under normal gravity conditions in the bioreactors, GS production was found to be unaffected by simulated microgravity. The repressive effect of glycerol in flask fermentations was not observed in the HARV. Thus the negative effect of glycerol on specific GS formation is dependent on shear and/or vessel geometry, not gravity.
47 citations
TL;DR: Rotating bioreactors designed at NASA’s Johnson Space Center were used to simulate a microgravity environment in which to study secondary metabolism and the degree of β-lactam antibiotic production was markedly inhibited by simulated microgravity.
Abstract: Rotating bioreactors designed at NASA's Johnson Space Center were used to simulate a microgravity environment in which to study secondary metabolism. The system examined was beta-lactam antibiotic production by Streptomyces clavuligerus. Both growth and beta-lactam production occurred in simulated microgravity. Stimulatory effects of phosphate and L-lysine, previously detected in normal gravity, also occurred in simulated microgravity. The degree of beta-lactam antibiotic production was markedly inhibited by simulated microgravity.
46 citations
TL;DR: Increasing ammonium concentrations added to chemically-defined medium MM enhanced the formation of LAT and decreased production of cephalosporins by S. clavuligerus and in cultures grown in the absence of lysine, greater enhancement of LAT formation by ammonium and less repression of cepsporin biosynthesis were observed.
Abstract: The L-lysine ɛ-aminotransferase (LAT) of Streptomyces clavuligerus was partially purified and characterized. The 51.3-kDa enzyme exhibited optimal activity at pH 7.0–7.5 and 30°C. It catalyzed transfer of the terminal amino group of L-lysine or L-ornithine to α -ketoglutarate. Oxalacetate and pyruvate were also used as acceptors of the amino group but with very low efficiency. Increasing ammonium concentrations added to chemically-defined medium MM enhanced the formation of LAT and decreased production of cephalosporins by S. clavuligerus. In cultures grown in the absence of lysine, greater enhancement of LAT formation by ammonium and less repression of cephalosporin biosynthesis were observed. In the chemically-defined GSPG medium, ammonium ions decreased cephalosporin production without showing an effect on LAT formation.
45 citations
TL;DR: Actinomadura sp.
Abstract: Actinomadura sp. strain 2966 can effectively convert compactin to pravastatin. The degree of conversion observed was 65% to 78% of compactin added and 65% to 88% of compactin taken up, depending on the concentration of compactin and duration of the experiment. Increasing the compactin concentration resulted in a higher final pravastatin concentration especially when compactin was added intermittently. Higher glucose concentrations had no effect on the bioconversion although uptake of compactin was inhibited. The conversion was linear over 16 hours. The system requires no induction and thus appears to be different from previously studied hydroxylases from actinomycetes.
17 citations
TL;DR: When oxygen limitation is used to decrease growth in a glucose-based chemically defined medium, specific MccB17 production to be positively related to growth rate and extent, and when various nutritional variations of media are examined, specific production of MCCB17 showed a negative relationship to growth rates and extent.
Abstract: Previous studies [Connell et al. (1987) Mol Microbiol 1: 195–201] have shown that expression of the microcin B17 (MccB17) promoter is inversely related to the growth rate of the culture, when slower growth was brought about by limitation of sources of carbon, nitrogen or phosphorus. When we used oxygen limitation to decrease growth in a glucose-based chemically defined medium, we found specific MccB17 production to be positively related to growth rate and extent. On the other hand, when we examined various nutritional variations of media, specific production of MccB17 showed a negative relationship to growth rate and extent, as would be predicted by the findings of Connell et al. (1987). Glucose, glycerol and acetate were found to repress MccB17 production; succinate was not repressive. Succinate is an excellent carbon source for production of MccB17 since high levels can be used with no or little interference in product synthesis.
14 citations
TL;DR: In this paper, Resting cells were crucial in elucidating the steps of vitamin B12 biosynthesis, and their use led to the following conclusions: (1) 5-aminolevulinic acid is a common precursor of vitamin b12 and porphyrins in various groups of microorganisms.
Abstract: Resting cells were crucial in elucidating the steps of vitamin B12 biosynthesis, and their use led to the following conclusions: (1) 5-aminolevulinic acid is a common precursor of vitamin B12 and porphyrins in various groups of microorganisms. (2) Glutamine is the donor of the amide groups in the biosynthesis of the amidated corrin structure of vitamin B12. (3) Methylated derivatives of uroporphyrinogen III, called corriphyrins, are precursors of the corrin ring of vitamin B12. They are formed in the initial steps of uroporphyrinogen III transformation to cobyrinic acid. (4) The initial common steps of heme and vitamin B12 synthesis are controlled by heme, while the biosynthetic pathway starting with methylation of uroporphyrinogen III and leading to corrinoids is controlled by vitamin B12. Aeration interferes with vitamin B12 formation but not that of porphyrins. The mechanism is that of oxygen suppression of methylation of uroporphyrinogen III. These conclusions were completely supported by subs...
11 citations
9 citations
TL;DR: The results thus show that the negative effects of 2% glycerol and 3% starch and 100 mM phosphate on cephalosporin production are not due to an effect on production of LAT activity, however, repression of Lat activity by 3% Glycerol would be expected to play a negative role in antibiotic production.
Abstract: Cephalosporin production by Streptomyces clavuligerus is known to be negatively regulated by carbon sources, e.g., glycerol and starch, and by phosphate at high concentrations. Formation of lysine ɛ-aminotransferase (LAT) activity, the first enzyme of the biosynthetic pathway, was affected by a high concentration of carbon source. Whereas 3% starch more than doubled LAT activity production as compared to 1% starch, 3% glycerol repressed LAT activity formation by 20%–30%. LAT activity production was not affected by 100 mM K2HPO4. Our results thus show that the negative effects of 2% glycerol and 3% starch and 100 mM phosphate on cephalosporin production are not due to an effect on production of LAT activity. However, repression of LAT activity by 3% glycerol would be expected to play a negative role in antibiotic production.
9 citations