Strategies for Production of Single Chain Antibody Fragment in E. Coli

ISBN-13: 9783659482892 / Angielski / Miękka / 2014 / 124 str.

Patil Ganesh

Opis

Recombinant expression of antibody fragments in E. coli is often hampered by low expression, inefficient translocation and aggregation in the periplasm. Aggregation problem during in vitro-renaturation is also a limiting factor, when antibody fragments expressed in bacterial cytosol as inclusion bodies. The objective of this work was to achieve and optimise periplasmic production, and to explore novel strategies for in vitro renaturation from cytoplasmic IBs. For the study, the single chain antibody fragment against the hapten oxazolone was chosen as the model system. In this book, results on periplasmic expression and expression as inclusion bodies and refolding thereof are presented. In expression in periplasmic effect of promoter strength and growth cultivation are presented. In vitro renaturation of IBs lead to establishment refolding protocols comprising of Ionic Liquids, which are emerging as green alternatives. Besides, we demonstrated usage of aromatic thiols as redox buffers components for protein refolding. Findings presented herein might be helpful for the of the periplasmic production of antibody fragments as as well as for establish protocols at physiological pH.

O autorze

Ganesh Patil has completed his Ph.D from Martin Luther University, Halle, Germany. His expertise lies in the area of recombinant protein cloning, expression, purification, characterization. He is specialized in the area of process development on production of recombinant proteins, bacterial and viral antigens and monoclonal antibodies.