A cost effective process development for enhanced expression of antibody fragment in microbial cells, its impact on cellular physiology and transcriptome

ABSTRACT: A process for manufacturing the recombinant Ranibizumab has been developed with overall productivity of 0.72±0.02 mg/ml. Initially this study was attempted to find critical medium components based on literature. Among all the additives, glutamine, proline, arginine and ethanol showed increase in protein production compared to control. There was approximately 2 fold increase in Ranibizumab production in all the four additives analyzed using RP HPLC. Addition of glutamine, arginine and proline amino acids resulted in enhanced protein production however, the pH of the medium changed variably (pH 3.5-6.0). Since economic considerations decides the longevity of the process, as it plays a major role when large-scale operations are considered. Therefore, after economical potential analysis ethanol was finalized as the medium additive. Later, its effect was checked on cell size (control 1.82 µm and optimized 2µm), cellular physiology (no cell swelling or shrinkage was observed, cell width and length came out almost same in both the cases) and cellular viability (was found even better in optimized). Later, RNAseq was also performed to find out the underlying mechanism. Based on the literature as well as transcriptomic analysis and RT-qPCR validation, changes in membrane properties and DNA synthesis resulting in gene amplification that may have enhanced the synthesis of inducible proteins in case of optimized medium. The transcriptome studies always generates a huge dataset that can act as valuable resource for future studies to perform host cell engineering with the aim of improving heterologous protein expression.

ORGANISM(S): Escherichia coli

PROVIDER: GSE143359 | GEO | 2023/01/09

REPOSITORIES: GEO