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  • Title: Two-cistron system overexpression of chloroplast glyceraldehyde-3-phosphate dehydrogenase subunit B and B-derivatives from spinach in Escherichia coli.
    Author: Tang GL, Wang YF, Bao JS, Chen HB.
    Journal: Protein Expr Purif; 2001 Jun; 22(1):31-7. PubMed ID: 11388796.
    Abstract:
    A gene coding for the subunit B (GapB) of chloroplast glyceraldehyde-3-phosphate dehydrogenase from spinach and its two derivatives (GapBc) lacking the GapB-specific C-terminal extension have been cloned by RT-PCR. These three genes have been overexpressed with full activity in Escherichia coli when a two-cistron expression system controlled by an inducible promoter P(trc) is used. With a suitable base composition of the first cistron, the expression level of GapB and the derivatives GapBc are expressed up to 15-20% of the total cell protein and around 20 mg of recombinant GapBcs with full activity are purified from 1 liter of cultured bacteria. The specific activity of the two derivatives GapBc (40-60 u/mg) is similar to that of GapA (50-70 u/mg) and lower than that of reported GapBc derivative (E. Baalmann, R. Scheibe, R. Cerff, and W. Martin, 1996, Plant Mol. Biol. 32, 505-513).
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