Secretory production and purification of functional full-length streptavidin from Bacillus subtilis.
Streptavidin is a versatile molecule for many in vitro and in vivo applications. To optimize the production of the full-length streptavidin in a soluble and functional form via secretion using Bacillus subtilis as the expression host, three different strategies were used. These strategies include the construction of a synthetic streptavidin gene, the installation of a transcription terminator, and the use of a sporulation mutant strain. In comparison with the wild-type streptavidin gene in expression studies, a combination of these approaches resulted in a 2.3-fold increase in streptavidin production. The production yields in complex and semidefined media were 94 and 24 mg/liter, respectively. A simple purification scheme which requires only a single ion-exchange matrix was designed to purify streptavidin to homogeneity directly from the culture supernatant. Purified streptavidin was in full length with good biotin binding capacity (3.2 binding sites available per tetramer). A combination of this expression system and purification scheme would be useful for production and purification of high-quality functional streptavidin for characterizations and practical applications.
Wu
Sau-Ching
SC
Department of Biological Sciences, University of Calgary, Calgary, Alberta T2N 1N4, Canada.
Hassan Qureshi
Mohammad
M
Wong
Sui-Lam
SL
D001412:Bacillus subtilis; D001710:Biotin / Q000378:metabolism; D005822:Genetic Vectors; D013058:Mass Spectrometry; D009154:Mutation; D019809:Streptavidin / Q000235:genetics* / Q000302:isolation & purification*
D001710:Biotin; D019809:Streptavidin