Control of formation of active soluble or inactive insoluble baker's yeast alpha-glucosidase PI in Escherichia coli by induction and growth conditions.
Using standard growth conditions (LB medium, 37 degrees C, induction with 5 mM IPTG) yeast alpha-glucosidase PI expressed under the control of the regulated tac-hybrid promoter results in the synthesis of insoluble aggregated alpha-glucosidase granules in Escherichia coli. Under these conditions active soluble alpha-glucosidase amounts to less than 1% of the heterologously produced protein. However, the amount of soluble active alpha-glucosidase was dramatically increased when the strong tac-hybrid promoter was to a limited extent induced. This was achieved at concentrations of 0.01 mM IPTG or of 1% lactose or lower in a lactose-permease deficient host strain containing the lacIq repressor gene on an R-plasmid. The formation of active soluble alpha-glucosidase was almost 100% when E. coli cells induced in this manner were cultivated under conditions that reduced growth rate, i.e. at decreased temperature, extreme pH values or in minimal and complete media supplemented with different carbon sources.
Kopetzki
E
E
Boehringer Mannheim GmbH, Department of Genetics, Penzberg, Federal Republic of Germany.
Schumacher
G
G
Buckel
P
P
D003470:Culture Media; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D005786:Gene Expression Regulation; D005800:Genes, Fungal; D006863:Hydrogen-Ion Concentration; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D012995:Solubility; D013696:Temperature; D000520:alpha-Glucosidases / Q000096:biosynthesis* / Q000235:genetics
D003470:Culture Media; D000520:alpha-Glucosidases