Engineered Sigma Factors Increase Full-Length Antibody Expression in Escherichia coli

Escherichia coli is one of the organisms of choice for production of antibody fragments such as Fabs and half-antibodies, however, obtaining high yields of full-length antibodies in E. coli remains a challenge. Global transcription machinery engineering (gTME), a powerful approach to improve cellular phenotypes, was used to improve expression of a full-length antibody (mAb1) in E. coli. A mutant library of one of the primary sigma factors in E. coli, RpoD (σ⁷⁰), was generated and cells expressing RpoD variants were screened for high levels of mAb1 expression using fluorescence assisted cell sorting (FACS). Three RpoD mutants were isolated via FACS and were shown to increase mAb1 expression by ~2–fold compared to the wild type RpoD. Transcriptional analysis using RNAseq of the cells expressing the sigma factor mutants revealed several interesting insights into the mechanistic role of these mutants in improving expression of mAb1.