Glutamate Dehydrogenase Specific Activity and Cephalosporin C Synthesis in the M8650 Series of Cephalosporium acremonium Mutants

Abstract

Conditions for the accurate measure of glutamic dehydrogenase (GDH) from Cephalosporium acremonium were determined. Km values for a-ketoglutarate and ammonium ion were 7 and 15 mM, respectively. The half-saturation for reduced nicotinamide adenine dinucleotide phosphate was 5 ,M. Reduced nicotinamide adenine dinucleotide did not serve as a cofactor for the enzyme. The specific activity of GDH was measured in six mutants of C. acremonium which varied in their ability to synthesize cephalosporin C. The mutants represented two separately derived lines, A and B. The four mutants in line B were characterized by a derepression of the GDH upon entry into stationary phase. The two mutants in line A were characterized by repressed levels of GDH during the same period. Both lines exhibited high GDH activity early in their fermentations, but activity decreased during the period of active cell growth. Cytochrome c concentrations followed the same pattern as total soluble intracellular protein. Line A mutants were low in cephalosporin C productivity and line B encompassed low, intermediate, and high productivity mutants. The relative frequency of yield improvements in line A and B indicate that the altered regulation pattern for GDH in line B may have removed a nitrogen limitation for cephalosporin C synthesis.

Enzymes involved in ammonia assimilation in the fungus Acremonium persicinum

“…Thus, Queener ct al. [22] determined the K m of NADP+-dependent ODH for 2-0xoglutarate and ammonia, as well as for NADPH in Cephalospotium acremonium. Their K m values obtained for 2-oxoglutarate and ammonia were 8 × 10 -3 M and 1.4 x 10 -2 M, respectively.…”

Strain Improvement and Preservation of β-Lactam-Producing Microorganisms

“…The Wisconsin group showed a correlation between increased productivity and reduced sporulation and growth QUEENER et al 1975). In the Wisconsin series, the greatest change was observed in the early ancestry.…”

“…In a study with improved mutants at Eli Lilly and Co., QUEENER et al (1975) reported that the specific activity of glutamate dehydrogenase was derepressed whereas two mutants in a low-yielding series had repressed levels of glutamate de-hydrogenase. The altered regulation pattern for this enzyme may have removed a nitrogen limitation for cephalosporin C synthesis.…”

Microbial Culture Selection

“…In a study on a series of improved cephalosporin C mutants, Queener et al (1975) reported that the specific activity of glutamate dehydrogenase (GDH) was derepressed upon entry into the stationary growth phase, whereas two mutants in a low-yielding series had repressed levels of 12. Microbial Culture Selection 293 GDH at the same growth phase.…”