Cytosolic Isobutanol Pathway Localization for the Production of Isobutanol

摘要

The present invention provides recombinant microorganisms comprising isobutanol producing metabolic pathway with at least one isobutanol pathway enzyme localized in the cytosol, wherein said recombinant microorganism is selected to produce isobutanol from a carbon source. Methods of using said recombinant microorganisms to produce isobutanol are also provided. In various aspects of the invention, the recombinant microorganisms may comprise a cytosolically active isobutanol pathway enzymes. In some embodiments, the invention provides mutated, modified, and/or chimeric isobutanol pathway enzymes with cytosolic activity. In various embodiments described herein, the recombinant microorganisms may be microorganisms of the Saccharomyces clade, Crabtree-negative yeast microorganisms, Crabtree-positive yeast microorganisms, post-WGD (whole genome duplication) yeast microorganisms, pre-WGD (whole genome duplication) yeast microorganisms, and non-fermenting yeast microorganisms.

主权项

1. A recombinant yeast microorganism comprising an isobutanol producing metabolic pathway, wherein said isobutanol producing metabolic pathway comprises the following substrate to product conversions:
(i) pyruvate to acetolactate; (ii) acetolactate to 2,3-dihydroxyisovalerate; (iii) 2,3-dihydroxyisovalerate to α-ketoisovalerate; (iv) α-ketoisovalerate to isobutyraldehyde; and (v) isobutyraldehyde to isobutanol;wherein
a) the enzyme that catalyzes a substrate to product conversion of pyruvate to acetolactate is an acetolactate synthase; b) the enzyme that catalyzes a substrate to product conversion of acetolactate to 2,3-dihydroxyisovalerate is a ketol-acid reductoisomerase derived from Lactococcus lactis; c) the enzyme that catalyzes a substrate to product conversion of 2,3-dihydroxyisovalerate to α-ketoisovalerate is a dihydroxy acid dehydratase; d) the enzyme that catalyzes a substrate to product conversion of α-ketoisovalerate to isobutyraldehyde is an α-ketoisovalerate decarboxylase; and e) the enzyme that catalyzes a substrate to product conversion of isobutyraldehyde to isobutanol is an alcohol dehydrogenase.