Optimization of the production and purification processes of carnobacteriocins Cbn BM1 and Cbn B2 from Carnobacterium maltaromaticum CP5 by heterologous expression in Escherichia coli.
An optimization of the production and purification processes of carnobacteriocins Cbn BM1 and Cbn B2 from Carnobacterium maltaromaticum CP5, by heterologous expression in Escherichia coli is described. The genes encoding mature bacteriocin were cloned into an E. coli expression system and expressed as a fusion protein with a thermostable thioredoxin. Recombinant E. coli were cultivated following a fed-batch fermentation process with pH, temperature and oxygenation regulation. The overexpression of the fusion proteins was improved by replacing IPTG by lactose. The fusion proteins were purified by thermal coagulation followed by affinity chromatography. The thioredoxin fusion protein was removed by using CNBr instead of enterokinase and the carnobacteriocins were recovered by reverse-phase chromatography. These optimizations led us to produce up to 320 mg of pure protein per liter of culture, which is four to ten fold higher than what is described for other heterologous expression systems.
Jasniewski
Jordane
J
Laboratoire de Science et Génie Alimentaires (LSGA), Nancy-Université, 2 avenue de la Forêt de Haye, B.P. 172 F-54505 Vandoeuvre-lès-Nancy, France.
Cailliez-Grimal
Catherine
C
Gelhaye
Eric
E
Revol-Junelles
Anne-Marie
AM
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D001430:Bacteriocins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D002846:Chromatography, Affinity; D003470:Culture Media / Q000737:chemistry; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005285:Fermentation; D015870:Gene Expression; D007544:Isopropyl Thiogalactoside / Q000378:metabolism; D007778:Lactobacillus / Q000235:genetics / Q000378:metabolism*; D007785:Lactose / Q000378:metabolism; D008969:Molecular Sequence Data; D015202:Protein Engineering; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013879:Thioredoxins / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D001430:Bacteriocins; D003470:Culture Media; D011993:Recombinant Fusion Proteins; D007544:Isopropyl Thiogalactoside; D013879:Thioredoxins; D007785:Lactose