Interplay of SOS induction, recombinant gene expression, and multimerization of plasmid vectors in Escherichia coli.
Using pBR322- and pUC-derived plasmid vectors, a homologous (Escherichia coli native esterase) and three heterologous proteins (human interleukin-2, human interleukin-6, and Zymomonas levansucrase) were synthesized in E. coli IC2015(recA::lacZ) and GY4786 (sfiA::lacZ) strains. Via time-course measurement of beta-galactosidase activity in each recombinant culture, the SOS induction was estimated in detail and the results were systematically compared. In recombinant E. coli, the SOS response did not happen either with the recombinant insert-negative plasmid backbone alone or the expression vectors containing the homologous gene. Irrespective of gene expression level and toxic activity of synthesized foreign proteins, the SOS response was induced only when the heterologous genes were expressed using a particular plasmid vector, indicating strong dependence on the recombinant gene clone and the selection of a plasmid vector system. It is suggested that in recombinant E. coli the SOS response (i.e., activation of recA expression and initial sfiA expression) may be related neither to metabolic burden nor toxic cellular event(s) by synthesized heterologous protein, but may be provoked by foreign gene-specific interaction between a foreign gene and a plasmid vector. Unlike in E. coli XL1-blue(recA(-)) strains used, all expression vectors encoding each of the three heterologous proteins were multimerized in E. coli IC2015 strains in the course of cultivation, whereas the expression vectors containing the homologous gene never formed the plasmid multimers. The extent of multimerization was also dependent on a foreign gene insert in the expression vector. As a dominant effect of the SOS induction, recombinant plasmid vectors used for heterologous protein expression appear to significantly form various multimers in the recA(+) E. coli host.
Lee
Jeewon
J
Department of Chemical and Biological Engineering, Korea University, 1,5 Ka, Anam-Dong, Seongbuk-Ku, Seoul 136-701, South Korea. leejw@korea.ac.kr
Kim
Hyung-Cheol
HC
Kim
Sung-Woo
SW
Kim
Seung Wook
SW
Hong
Suk In
SI
Park
Young-Hoon
YH
D002460:Cell Line; D004926:Escherichia coli / Q000145:classification / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D005822:Genetic Vectors; D010957:Plasmids; D011938:Rec A Recombinases / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D011995:Recombination, Genetic; D013014:SOS Response, Genetics; D012680:Sensitivity and Specificity; D013045:Species Specificity; D001616:beta-Galactosidase / Q000235:genetics
D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; C053139:sulA protein, E coli; D011938:Rec A Recombinases; D001616:beta-Galactosidase