Application of factorial and Doehlert designs for optimization of pectate lyase production by a recombinant Escherichia coli |
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Authors: | C G Hounsa J M Aubry H C Dubourguier J P Hornez |
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Institution: | (1) Laboratoire de Microbiologie Fondamentale et Appliquée, Université des Sciences et Technologies de Lille, Batiment SN2, F-59655 Villeneuve d’Ascq, France. Fax: 33-20436504, FR;(2) Laboratoire de Chimie de Formulation, Ecole Supérieure de Chimie de Lille, B.P. 108, 59652 Villeneuve d’Ascq Cedex, France, FR |
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Abstract: | The expression of a recombinant pectate lyase from Bacteroides thetaiotaomicron strain 217 was studied in Escherichia coli strain HB101(pBT4). First, two sets of complete 24 factorial designs were used to evaluate the influences of casamino acids, glucose, magnesium, calcium, tetracycline, ampicillin,
tryptophan and MOPS buffer on pectate lyase production in a basal medium. While casamino acids, glucose and magnesium were
found to be the prevalent factors, the presence of tetracycline, ampicillin and MOPS buffer were necessary for the reproducibility
of the process, probably by increasing the plasmid stability. Secondly, application of the Doehlert design, a response-surface
methodology, allowed a good prediction of pectate lyase production according to the variation in glucose and magnesium concentrations.
This optimization strategy allowed the production of biomass and recombinant pectate lyase respectively to be increased from
0.2 g l-1 to 1.9 g l-1 (dry weight) and from 10 units ml-1 to 210 units ml-1 within 24 h at 30°C in shake flasks.
Received: 26 July 1995/Received revision: 22 January 1996/Accepted: 29 January 1996 |
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