Screening for conditions of enhanced production of a recombinant beta-glucanase secreted into the medium by Escherichia coli.
The extracellular production of a hybrid bacterial beta-glucanase using Escherichia coli was studied by using combinations of promoters of varying strength for both a beta-glucanase as the target protein and the Kil protein as the releasing factor. Four strains with different combinations of promoter strengths were cultivated in shake-flasks on four different media to assess the cross-influence of promoter and medium in a general manner. Promoters were taken from natural as well as synthetic sequences known to exhibit either weak or strong promoter strength. By far the highest extracellular glucanase activity (>200 U ml(-1)) was achieved when a strain harbouring the kil gene under control of a strong synthetic stationary-phase promoter and the glucanase gene under control of a strong synthetic constitutive promoter was cultivated on a complex medium mainly composed of casein peptone, yeast extract, and glycerol.
Spexard
Meike
M
Fermentation Engineering, Faculty of Technology, Bielefeld University, 33594 Bielefeld, Germany.
Beshay
Usama
U
Risse
Joe Max
JM
Miksch
Gerhard
G
Flaschel
Erwin
E
D001681:Biological Assay / Q000379:methods*; D044644:Endo-1,3(4)-beta-Glucanase / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000502:physiology*; D024861:Genetic Enhancement / Q000379:methods*; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000378:metabolism*
D011994:Recombinant Proteins; D044644:Endo-1,3(4)-beta-Glucanase