Peroxisomes and peroxisomal transketolase and transaldolase enzymes are essential for xylose alcoholic fermentation by the methylotrophic thermotolerant yeast, Ogataea (Hansenula) polymorpha

• Published in: Biotechnology for biofuels Vol. 11; no. 1; p. 197
• Main Authors: Kurylenko, Olena O, Ruchala, Justyna, Vasylyshyn, Roksolana V, Stasyk, Oleh V, Dmytruk, Olena V, Dmytruk, Kostyantyn V, Sibirny, Andriy A
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 19.07.2018; BioMed Central; BMC
• Subjects: Amino acids; Ammonium; Analysis; Biosynthesis; Cloning; Conditional mutant; Dehydrogenases; Dihydroxyacetone; Dihydroxyacetone synthase; Drug resistance; Enzymes; Ethanol; Fermentation; Fungi; Glucose; Hansenula polymorpha; High temperature; High-temperature alcoholic fermentation; Metabolic engineering; Metabolism; Methylotrophs; Mutants; Nitrate reductase; Ogataea (Hansenula) polymorpha; Ogataea polymorpha; Pentose; Peroxisomes; Pichia stipitis; Scheffersomyces stipitis; Sugar; Temperature; Transaldolase; Transketolase; Xylose; Yeast; Xylose; Peroxisomes; Ethanol; Ogataea (Hansenula) polymorpha; High-temperature alcoholic fermentation
• ISSN: 1754-6834; 1754-6834
• DOI: 10.1186/s13068-018-1203-z

is one of the most thermotolerant xylose-fermenting yeast species reported to date. Several metabolic engineering approaches have been successfully demonstrated to improve high-temperature alcoholic fermentation by . Further improvement of ethanol production from xylose in depends on the identification of bottlenecks in the xylose conversion pathway to ethanol. Involvement of peroxisomal enzymes in xylose metabolism has not been described to date. Here, we found that peroxisomal transketolase (known also as dihydroxyacetone synthase) and peroxisomal transaldolase (enzyme with unknown function) in the thermotolerant methylotrophic yeast, , are required for xylose alcoholic fermentation, but not for growth on this pentose sugar. Mutants with knockout of and coding for peroxisomal transketolase and peroxisomal transaldolase, respectively, normally grow on xylose. However, these mutants were found to be unable to support ethanol production. The mutant with the knockout (coding for cytosolic transaldolase) normally grew on glucose and did not grow on xylose; this defect was rescued by overexpression of . The conditional mutant, - , that expresses the cytosolic transketolase gene under control of the ammonium repressible nitrate reductase promoter did not grow on xylose and grew poorly on glucose media supplemented with ammonium. Overexpression of only partially restored the defects displayed by the - mutant. The mutants defective in peroxisome biogenesis, and , showed normal growth on xylose, but were unable to ferment this sugar. Moreover, the mutant of the non-methylotrophic yeast, , normally grows on and ferments xylose. Separate overexpression or co-overexpression of and in the wild-type strain increased ethanol synthesis from xylose 2 to 4 times with no effect on the alcoholic fermentation of glucose. Overexpression of and also elevated ethanol production from xylose. Finally, co-overexpression of and in the best previously isolated xylose to ethanol producer led to increase in ethanol accumulation up to 16.5 g/L at 45 °C; or 30-40 times more ethanol than is produced by the wild-type strain. Our results indicate the importance of the peroxisomal enzymes, transketolase (dihydroxyacetone synthase, Das1), and transaldolase (Tal2), in the xylose alcoholic fermentation of .