Abstract
The protein composition of inclusion bodies produced in recombinant Escherichia coli overproducing Vitreoscilla hemoglobin (VHb) was analyzed by one-dimensional and two-dimensional electrophoresis techniques. Results indicate the presence of two types of cytoplasmic aggregates of differing morphology in single bacterial cells. These aggregates also differ in their relative content of VHb and pre-beta-lactamase and are separable by differential centrifugation. Results further suggest that the cytoplasmic protein elongation factor Tu is integrated into VHb inclusion bodies. The presence of the outer membrane proteins OmpA and OmpF in inclusion body preparations is attributed to cell envelope contamination rather than specific involvement in inclusion bodies. The specificity of in vivo protein aggregation is discussed.
Highlights
A large number of highly diverse proteins have been insolubilized in this way, and it, appears that the phenomenon of recombinant protein inclusion body formation is not unique to E. coli or, for that matter, to bacterial cells (Kitano et al, 1987a)
Our results indicate the presence of two types of cytoplasmic inclusion bodies differing in their relative content of Vitreoscilla hemoglobin (VHb) and pre-glactamase, both expressed from genes carried on the same plasmid
Microscopic analysis indicates that two types of inclusion bodies, differing in general morphology, are produced in single cells of JM101:pREDZ
Summary
Results indicate the presence of two types of cytoplasmic aggregates of differing morphology in single bacterial cells. These aggregates differ in their relative content of Vl3b and pre+-lactamase and are separable by differential centrifugation. The expression of a recombinant protein in the bacterial cell Eschrichia coli is often accompanied by the insolubilization of that protein in viva into aggregates referred to as inclusion bodies. A large number of highly diverse proteins have been insolubilized in this way, and it, appears that the phenomenon of recombinant protein inclusion body formation is not unique to E. coli or, for that matter, to bacterial cells (Kitano et al, 1987a). Several incisive reviews of the literature concerning inclusion body formation have been prepared (Kane and Hartley, 1988; Mitraki and King, 1989; Schein, 1989)
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