Conversion of an inactive xylose isomerase into a functional enzyme by co-expression of GroEL-GroES chaperonins in Saccharomyces cerevisiae

Abstract

BackgroundSecond-generation ethanol production is a clean bioenergy source with potential to mitigate fossil fuel emissions. The engineering of Saccharomyces cerevisiae for xylose utilization is an essential step towards the production of this biofuel. Though xylose isomerase (XI) is the key enzyme for xylose conversion, almost half of the XI genes are not functional when expressed in S. cerevisiae. To date, protein misfolding is the most plausible hypothesis to explain this phenomenon.ResultsThis study demonstrated that XI from the bacterium Propionibacterium acidipropionici becomes functional in S. cerevisiae when co-expressed with GroEL-GroES chaperonin complex from Escherichia coli. The developed strain BTY34, harboring the chaperonin complex, is able to efficiently convert xylose to ethanol with a yield of 0.44 g ethanol/g xylose. Furthermore, the BTY34 strain presents a xylose consumption rate similar to those observed for strains carrying the widely used XI from the fungus Orpinomyces sp. In addition, the tetrameric XI structure from P. acidipropionici showed an elevated number of hydrophobic amino acid residues on the surface of protein when compared to XI commonly expressed in S. cerevisiae.ConclusionsBased on our results, we elaborate an extensive discussion concerning the uncertainties that surround heterologous expression of xylose isomerases in S. cerevisiae. Probably, a correct folding promoted by GroEL-GroES could solve some issues regarding a limited or absent XI activity in S. cerevisiae. The strains developed in this work have promising industrial characteristics, and the designed strategy could be an interesting approach to overcome the non-functionality of bacterial protein expression in yeasts.Electronic supplementary materialThe online version of this article (10.1186/s12896-017-0389-7) contains supplementary material, which is available to authorized users.

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“…Numerous GroEL (60 kDa) and GroES (10 kDa) chaperonins were detected and assigned, amongst other taxa, to Clostridia and Bacteroidia ( Supplementary Table S3 ). Recently, the co-expression of GroEL-GroES was found to be imperative for the production of a functional xylose isomerase in Saccharomyces cerevisiae ( Temer et al, 2017 ). Xylose isomerases were detected in both bioreactor fractions and assigned exclusively to Clostridia and Bacteroidia ( Supplementary Table S3 ).…”

Optimizing anaerobic growth rate and fermentation kinetics in Saccharomyces cerevisiae strains expressing Calvin-cycle enzymes for improved ethanol yield

“…Functional expression of a plant RuBisCO in E. coli was also recently shown to require co-expression of no fewer than five plant chaperones [ 74 ], highlighting the importance of expression of folding-assisting proteins in the formation of functional RubisCO complexes. In the case of GroEL and GroeS specifically, it was recently shown that their expression facilitates functional expression of several heterologous proteins in yeasts [ 28 , 75 , 76 ]. Further, their expression can potentially be beneficial in improving strain robustness against industrial fermentation conditions [ 77 ].…”

Construction of advanced producers of first- and second-generation ethanol in Saccharomyces cerevisiae and selected species of non-conventional yeasts (Scheffersomyces stipitis, Ogataea polymorpha)

“…Expression of the codon-optimized XI of C. phytofermentans in S. cerevisiae resulted in a 46% increase in specific growth rate on xylose as compared to the strain expressing a non-optimized version of the gene [20]. XI from the bacterium Propionibacterium acidipropionici became functionally active in S. cerevisiae when coexpressed with GroEL-GroES chaperonin complex from E. coli [178]. More details on functional expression if XI in S. cerevisiae described in a recent reviews [72,103].…”