Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.637709
Title: The production of glutaminase from Aspergillus awamori
Author: Jones, F. N. H.
Awarding Body: University College of Swansea
Current Institution: Swansea University
Date of Award: 1995
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Abstract:
Using Aspergillus awamori ATCC 14335 in flask cultures, the growth medium was optimised for the production of extracellular glutaminase. The medium consisted of simple salts with glucose and soybean flour. Factors controlling the expression of glutaminase were the glucose : soybean flour ratio; the lower the ratio, the higher the levels of glutaminase. High levels of ammonia completely inhibited glutaminase production. The levels of magnesium sulphate and phosphate also influence production. Using these methods, initial activities of less than 200 nMoles/min/L were increased to 2000 nMoles/min/L in optimal conditions. The glutaminase was found to be primarily an intracellular enzyme and extracellular activity is a leakage product of the cell. Studies of disruption revealed ratios of intracellular to extracellular activities of 20:1. During growth in large scale cultures, this ratio was found to vary from under 5:1 to over 20:1. Using 5 and 50 litre bioreactors the scale-up and production of glutaminase was investigated. Studies of the time course of the fermentations for glutaminase production supported the control mechanisms inferred in the shake flask studies. Intracellular glutaminase production peaked in a narrow window between glucose depletion and high concentrations of ammonia in the growth medium. Extracellular activities were detected after the intracellular production had peaked. Glutaminase levels varied by at least a factor of two between sub-optimal and optimal harvesting conditions (i.e. intracellular activities of 7000-18000 nMoles/min/L). Intensification of the fermentation was limited by the levels of biomass which could be grown in the bioreactor. This was mainly due to the poor mixing associated with high concentrations of mycelia. Using a crude enzyme preparation, the enzyme was characterised and compared with enzymes in the literature. This showed that the enzyme was substantially more stable than those previously published e.g. half life (141 minutes at 40°C for the intracellular enzyme), Km values for the intracellular/extracellular enzymes were 0.432 nM and 0.330 mM respectively.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.637709  DOI: Not available
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