Fed-batch production of recombinant human calcitonin precursor fusion protein using Staphylococcus carnosus as an expression-secretion system.
A pH-auxostatic fed-batch process was developed for the secretory production of a fusion protein consisting of the pro-part of Staphylococcus hyicus lipase and two synthetic human calcitonin (hCT) precursor repeats under the control of a xylose-inducible promotor from Staphylococcus xylosus. Using glycerol as the energy source and pH-controlled addition of yeast extract resulted in the production of 2000 mg 1(-1) of the fusion protein (420 mg 1(-1) of the recombinant hCT precursor) within 14 h, reaching 45 g 1(-1) cell dry mass with Staphylococcus carnosus in a stirred-tank reactor. Product titer and space-time yield (30 mg calcitonin precursor 1(-1) h(-1)) were thus improved by a factor of 2, and 4.5, respectively, compared to Escherichia coli expression-secretion systems for the production of calcitonin precursors. Two hundred grams of the fusion protein was secreted by the recombinant S. carnosus on a 150-1 scale (scale-up factor of 50) with a minimum use of technical-grade yeast extract (40 mg fusion protein g(-1) yeast extract).
Dilsen
S
S
Lehrstuhl für Bioverfahrenstechnik, Technische Universität München, Garching, Germany.
Paul
W
W
Sandgathe
A
A
Tippe
D
D
Freudl
R
R
Thömmes
J
J
Kula
M R
MR
Takors
R
R
Wandrey
C
C
Weuster-Botz
D
D
D000596:Amino Acids / Q000378:metabolism; D019149:Bioreactors; D002116:Calcitonin / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D003470:Culture Media; D005285:Fermentation; D005990:Glycerol / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000235:genetics; D011401:Promoter Regions, Genetic; D011498:Protein Precursors / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000378:metabolism; D013210:Staphylococcus / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism
D000596:Amino Acids; D003470:Culture Media; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D002116:Calcitonin; D008049:Lipase; D005990:Glycerol