Proteome and transcriptome based analysis of Bacillus subtilis cells overproducing an insoluble heterologous protein

Bacillus subtilis and related Bacillus species are frequently used as hosts for the industrial production of recombinant proteins. In this study the c ellular response of B. subtilis to the overproduction of an insoluble heter ologous protein was investigated. For this purpose PorA, an outer membrane protein from Neisseria meningitidis, which accumulates after overexpression in the cytoplasm of B. subtilis mainly in the form of inclusion bodies, wa s used. The molecular response to overexpression of porA has been analysed at the transcriptional level using the DNA macro array technique and at the translational level by two-dimensional polyacrylamide gel electrophoresis. It was found that the expression of the heat shock genes of class I (dnaK, groEL and grpE) and class m (c1pP and c1pC) are increased under overproduc ing conditions. Furthermore, the protein levels of the two ribosomal protei ns RpsB and Rp1J are increased in the PorA overproducing cells. The transcr iptome analysis indicated that mRNA levels of genes encoding pyrimidine and purine synthesis enzymes but also from ribosomal protein genes have elevat ed levels under overproducing conditions. Finally, the association of the p rotease C1pP and its ATPase subunits C1pC and C1pX with the PorA inclusion bodies was demonstrated by means of the immunogold labelling technique.