Metabolic Engineering Kluyveromyces marxianus for Isoprene Production from Eucalyptus globulus Wood Cellulosic Fraction

Abstract

Isoprene, a highly volatile hydrocarbon with numerous industrial applications, has traditionally been produced from petrochemical sources through processes associated with significant environmental impacts. Microbial production of isoprene has emerged as a promising and more sustainable alternative. In this study, the potential of the non-conventional yeast Kluyveromyces marxianus to produce isoprene from a renewable feedstock was explored, contributing to the development of a more sustainable process. K. marxianus was engineered to produce isoprene from glucose through the expression of an isoprene synthase gene, and the gene copy number of this enzyme was found to significantly influence isoprene production. Furthermore, enhancing the supply of the isoprene precursors acetyl-CoA and dimethylallyl diphosphate (DMAPP) via engineering of the mevalonate pathway led to increased production. A higher headspace-to-culture ratio in sealed serum bottles also facilitated isoprene accumulation. Importantly, isoprene production was achieved from the cellulosic fraction of pretreated Eucalyptus globulus wood. To our knowledge, this is the first report of isoprene production in K. marxianus using a lignocellulosic feedstock, providing proof of concept for its potential in integrated processes based on sustainable substrates under stressful conditions.