scholarly journals Alleviation of Proteolytic Sensitivity To Enhance Recombinant Lipase Production in Escherichia coli

2009 ◽  
Vol 75 (16) ◽  
pp. 5424-5427 ◽  
Author(s):  
Niju Narayanan ◽  
C. Perry Chou
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Double Mutant ◽  
Protein Production ◽  
Functional Expression ◽  
Lipase Production ◽  
Recombinant Lipase ◽  
Pseudozyma Antarctica

ABSTRACT Two amino acids, Leu149 and Val223, were identified as proteolytically sensitive when Pseudozyma antarctica lipase (PalB) was heterologously expressed in Escherichia coli. The functional expression was enhanced using the double mutant for cultivation. However, the recombinant protein production was still limited by PalB misfolding, which was resolved by DsbA coexpression.

scholarly journals Decoupling of recombinant protein production from Escherichia coli cell growth enhances functional expression of plant Leloir glycosyltransferases

2019 ◽  
Vol 116 (6) ◽  
pp. 1259-1268 ◽  
Author(s):  
Martin Lemmerer ◽  
Juergen Mairhofer ◽  
Alexander Lepak ◽  
Karin Longus ◽  
Rainer Hahn ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Cell Growth ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Functional Expression ◽  
Coli Cell

scholarly journals Predictive approaches to guide the expression of recombinant vaccine targets in Escherichia coli: a case study presentation utilising Absynth Biologics Ltd. proprietary Clostridium difficile vaccine antigens

2021 ◽  
Author(s):  
Hirra Hussain ◽  
Edward A McKenzie ◽  
Andrew M Robinson ◽  
Neill A Gingles ◽  
Fiona Marston ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Clostridium Difficile ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Expression Systems ◽  
Predictive Approaches ◽  
Bacterial Expression Systems

AbstractBacterial expression systems remain a widely used host for recombinant protein production. However, overexpression of recombinant target proteins in bacterial systems such as Escherichia coli can result in poor solubility and the formation of insoluble aggregates. As a consequence, numerous strategies or alternative engineering approaches have been employed to increase recombinant protein production. In this case study, we present the strategies used to increase the recombinant production and solubility of ‘difficult-to-express’ bacterial antigens, termed Ant2 and Ant3, from Absynth Biologics Ltd.’s Clostridium difficile vaccine programme. Single recombinant antigens (Ant2 and Ant3) and fusion proteins (Ant2-3 and Ant3-2) formed insoluble aggregates (inclusion bodies) when overexpressed in bacterial cells. Further, proteolytic cleavage of Ant2-3 was observed. Optimisation of culture conditions and changes to the construct design to include N-terminal solubility tags did not improve antigen solubility. However, screening of different buffer/additives showed that the addition of 1–15 mM dithiothreitol alone decreased the formation of insoluble aggregates and improved the stability of both Ant2 and Ant3. Structural models were generated for Ant2 and Ant3, and solubility-based prediction tools were employed to determine the role of hydrophobicity and charge on protein production. The results showed that a large non-polar region (containing hydrophobic amino acids) was detected on the surface of Ant2 structures, whereas positively charged regions (containing lysine and arginine amino acids) were observed for Ant3, both of which were associated with poor protein solubility. We present a guide of strategies and predictive approaches that aim to guide the construct design, prior to expression studies, to define and engineer sequences/structures that could lead to increased expression and stability of single and potentially multi-domain (or fusion) antigens in bacterial expression systems.

scholarly journals Isolating Escherichia coli strains for recombinant protein production

2016 ◽  
Vol 74 (5) ◽  
pp. 891-908 ◽  
Author(s):  
Susan Schlegel ◽  
Pierre Genevaux ◽  
Jan-Willem de Gier
Keyword(s):  
Escherichia Coli ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Protein Production

scholarly journals Recombinant protein production in an Escherichia coli reduced genome strain

2007 ◽  
Vol 9 (2) ◽  
pp. 133-141 ◽  
Author(s):  
Shamik S. Sharma ◽  
Frederick R. Blattner ◽  
Sarah W. Harcum
Keyword(s):  
Escherichia Coli ◽  
Recombinant Protein ◽  
Recombinant Protein Production ◽  
Protein Production ◽  
Reduced Genome
Sign in / Sign up

Export Citation Format

Share Document