New strategies of protein engineering—directed evolution of enzymein vitro

1999 ◽  
Vol 44 (18) ◽  
pp. 1641-1648 ◽  
Author(s):  
Hongying Zhang ◽  
Xiangduo Kong ◽  
Jin Zhang
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
New Strategies

scholarly journals Protein Engineering Approaches to Enhance Fungal Laccase Production in S. cerevisiae

2021 ◽  
Vol 22 (3) ◽  
pp. 1157
Author(s):  
Pablo Aza ◽  
Felipe de Salas ◽  
Gonzalo Molpeceres ◽  
David Rodríguez-Escribano ◽  
Iñigo de la Fuente ◽  
...  
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
Signal Peptide ◽  
Laccase Activity ◽  
Laccase Production ◽  
Wide Range ◽  
Mating Factor ◽  
Conserved Amino Acids ◽  
Basidiomycete Fungi

Laccases secreted by saprotrophic basidiomycete fungi are versatile biocatalysts able to oxidize a wide range of aromatic compounds using oxygen as the sole requirement. Saccharomyces cerevisiae is a preferred host for engineering fungal laccases. To assist the difficult secretion of active enzymes by yeast, the native signal peptide is usually replaced by the preproleader of S. cerevisiae alfa mating factor (MFα1). However, in most cases, only basal enzyme levels are obtained. During directed evolution in S. cerevisiae of laccases fused to the α-factor preproleader, we demonstrated that mutations accumulated in the signal peptide notably raised enzyme secretion. Here we describe different protein engineering approaches carried out to enhance the laccase activity detected in the liquid extracts of S. cerevisiae cultures. We demonstrate the improved secretion of native and engineered laccases by using the fittest mutated α-factor preproleader obtained through successive laccase evolution campaigns in our lab. Special attention is also paid to the role of protein N-glycosylation in laccase production and properties, and to the introduction of conserved amino acids through consensus design enabling the expression of certain laccases otherwise not produced by the yeast. Finally, we revise the contribution of mutations accumulated in laccase coding sequence (CDS) during previous directed evolution campaigns that facilitate enzyme production.

Continuous directed evolution for strain and protein engineering

2018 ◽  
Vol 53 ◽  
pp. 158-163 ◽  
Author(s):  
Simon d’Oelsnitz ◽  
Andrew Ellington
Keyword(s):  
Protein Engineering ◽  
Directed Evolution

“Itching” for new strategies in protein engineering

10.1038/70699 ◽  
1999 ◽  
Vol 17 (12) ◽  
pp. 1159-1160 ◽  
Author(s):  
Stephen W. Michnick ◽  
Frances H. Arnold
Keyword(s):  
Protein Engineering ◽  
New Strategies

Protein Engineering of Carboxyl Esterases by Rational Design and Directed Evolution

2009 ◽  
Vol 16 (10) ◽  
pp. 1162-1171 ◽  
Author(s):  
U.T. Bornscheuer ◽  
D. Bottcher ◽  
M. Schmidt
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
Rational Design

A completely in vitro ultrahigh-throughput droplet-based microfluidic screening system for protein engineering and directed evolution

Lab on a Chip ◽  
2012 ◽  
Vol 12 (5) ◽  
pp. 882 ◽  
Author(s):  
Ali Fallah-Araghi ◽  
Jean-Christophe Baret ◽  
Michael Ryckelynck ◽  
Andrew D. Griffiths
Keyword(s):  
Protein Engineering ◽  
Directed Evolution ◽  
Screening System
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