REGULATION OF SCD4-183 GENE-EXPRESSION FROM PHAGE-T7-BASED VECTORS INESCHERICHIA-COLI

In this paper, we describe various parameters affecting the regulation of expression of the sCD4-183 gene, encoding the 183-amino-acid solub le human two-domain CD4 protein, from phage-T7-based pET vectors. We d emonstrated that for the sCD4-183 protein, the highest protein yield w as obtained using vector pET-9a, in which neither expression of the T7 RNA polymerase-encoding gene nor the target gene was tightly regulate d. The highest overall protein yield was obtained from cells grown for 24 h in the absence of inducer, a strategy that may be generally usef ul for production of less toxic proteins. We also describe two modific ations of the pET vector system that effectively minimized leaky (unin duced) expression and enhanced plasmid stability. These have potential use in the production of toxic proteins, or of non-toxic proteins pro duced in high-density cultures.