Rational strain engineering interventions to enhance cellulase secretion by Saccharomyces cerevisiae

Abstract

Lignocellulosic biomass remains an attractive feedstock for the production of fuels if a technology can be developed to overcome its recalcitrance. Consolidated bioprocessing (CBP) is one technology under development that aims to make this conversion process economically feasible. While no ideal CBP organism has been developed, several options have been pursued including engineering of the ethanologenic yeast Saccharomyces cerevisiae. Considering the genetic malleability of this model organism, a variety of chemicals and chemical precursors could also be produced directly from cellulosic feedstocks, assuming an enzymatic system for the hydrolysis of the feedstock sugar polymers can be established. While there have been several accounts of the secretion of cellulases by strains of S. cerevisiae and the successful conversion of limited amounts of amorphous and model crystalline cellulose feedstocks to ethanol, substantial conversion of crystalline cellulose by these strains in the absence of exogenous cellulases has not been reported. The most cited reasons for this were the low secretion titer of cellulases in general and of cellobiohydrolases in particular. This review will compare the efforts that have been made to enhance heterologous protein secretion in the yeast S. cerevisiae through rational strain engineering with a focus on cellulases and will investigate important factors in developing successful CBP-yeast strains.

Adapting the yeast consolidated bioprocessing paradigm for biorefineries

“…However, low secretion titres of cellulases lead to poor substrate hydrolysis efficiency and slow conversion rates, preventing their commercial application. Strategies since employed to improve secretion include engineering of peptide leader sequences, optimization of gene copy number, manipulation of promoter strength, and engineering the heterologous protein for codon optimization or to remove inhibition (Kroukamp et al, 2017). Rational strain improvements, including increasing ER-resident chaperones, accelerating vesicle fusion events, altering protein glycosylation, modulating cellular stress.…”

Rapid optimisation of cellulolytic enzymes ratios in Saccharomyces cerevisiae using in vitro SCRaMbLE

“…There have been many rational engineering approaches to enhance cellulase production and secretion in S. cerevisiae (reviewed in [ 15 ]), however, since the best gene ratio to produce efficient hydrolysis for each substrate is not always known, techniques that generate large libraries of DNA and/or strains containing different gene ratios provide a promising approach. In previous work, Yamada et al [ 16 ] produced a library of cellulolytic yeast strains using cocktail δ-integration.…”

Rapid optimisation of cellulolytic enzymes ratios in Saccharomyces cerevisiae using in vitro SCRaMbLE

“…There have been many rational engineering approaches to enhance cellulase production and secretion in S. cerevisiae (reviewed in [15]), however, since the best gene ratio to produce e cient hydrolysis for each substrate is not always known, techniques that generate large libraries of DNA and/or strains containing different gene ratios provide a promising approach. In previous work, Yamada et al [16] produced a library of cellulolytic yeast strains using cocktail δ-integration.…”