US 12,270,034 B2
Mut- methylotrophic yeast
Domen Zavec, Vienna (AT); Brigitte Gasser, Vienna (AT); and Diethard Mattanovich, Vienna (AT)
Assigned to UNIVERSITÄT FÜR BODENKULTUR WIEN, Vienna (AT)
Appl. No. 17/601,045
Filed by Universität für Bodenkultur Wien, Vienna (AT)
PCT Filed Apr. 1, 2020, PCT No. PCT/EP2020/059284
§ 371(c)(1), (2) Date Oct. 1, 2021,
PCT Pub. No. WO2020/201369, PCT Pub. Date Oct. 8, 2020.
Claims priority of application No. PCT/EP2019/058190 (WO), filed on Apr. 1, 2019; and application No. PCT/EP2019/058191 (WO), filed on Apr. 1, 2019.
Prior Publication US 2022/0170032 A1, Jun. 2, 2022
Int. Cl. C12N 15/81 (2006.01); C07K 14/765 (2006.01); C07K 16/06 (2006.01); C12N 9/04 (2006.01)
CPC C12N 15/815 (2013.01) [C07K 14/765 (2013.01); C07K 16/06 (2013.01); C12N 9/0006 (2013.01); C12Y 101/01001 (2013.01); C12Y 101/03013 (2013.01); C07K 2317/14 (2013.01); C07K 2317/569 (2013.01); C12N 2510/02 (2013.01); C12N 2830/002 (2013.01)] 25 Claims
 
1. A recombinant methanol utilization pathway deficient methylotrophic yeast (Mut−) host cell which is engineered:
a) by one or more genetic modifications to reduce expression of a first and a second endogenous gene compared to the host cell prior to said one or more genetic modifications, wherein
i. the first endogenous gene encodes alcohol oxidase 1 (Aox1) comprising the amino acid sequence identified as SEQ ID NO:1 or a homologue thereof, and
ii. the second endogenous gene encodes alcohol oxidase 2 (Aox2) comprising the amino acid sequence identified as SEQ ID NO:3 or a homologue thereof,
and
b) by one or more genetic modifications that increases expression of an alcohol dehydrogenase 2 (Adh2) gene compared to the host cell prior to said one or more genetic modifications, wherein the Adh2 gene encodes an alcohol dehydrogenase 2 (ADH2), and wherein the methylotrophic yeast is a P. pastoris strain.