Production of interferon-alpha in high cell density cultures of recombinant Escherichia coli and its single step purification from refolded inclusionbody proteins
Kr. Babu et al., Production of interferon-alpha in high cell density cultures of recombinant Escherichia coli and its single step purification from refolded inclusionbody proteins, APPL MICR B, 53(6), 2000, pp. 655-660
Escherichia coli TG1 transformed with a temperature-regulated interferon-al
pha expression vector was grown to high cell density in defined medium cont
aining glucose as the sole carbon and energy source, utilizing a simple fed
-batch process. Feeding was carried out to achieve an exponential increase
in biomass at growth rates which minimized acetate production. Thermal indu
ction of such high cell density cultures resulted in the production of simi
lar to 4 g interferon-alpha/l culture broth. Interferon-a was produced excl
usively in the form of insoluble inclusion bodies and was solubilized under
denaturing conditions, refolded in the presence of arginine and purified t
o near homogeneity, utilizing single-step ion-exchange chromatography on Q-
Sepharose. The yield of purified interferon-alpha was similar to 300 mg/l w
ith respect to the original high cell density culture broth (overall yield
of similar to 7.5% active interferon-alpha). The purified recombinant inter
feron-alpha was found by different criteria to be predominantly monomeric a
nd possessed a specific bioactivity of similar to 2.5 x 10(8) IU/mg based o
n viral cytopathic assay.