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Identification of genetic variants of the industrial yeast Komagataella phaffii (Pichia pastoris) that contribute to increased yields of secreted heterologous proteins

2022; Public Library of Science; Volume: 20; Issue: 12 Linguagem: Inglês

10.1371/journal.pbio.3001877

1545-7885

Benjamin Offei, Stephanie Braun‐Galleani, Anjan Venkatesh, William Casey, Kevin E. O’Connor, Kevin P. Byrne, Kenneth H. Wolfe,

Plant Reproductive Biology

The yeast Komagataella phaffii (formerly called Pichia pastoris ) is used widely as a host for secretion of heterologous proteins, but only a few isolates of this species exist and all the commonly used expression systems are derived from a single genetic background, CBS7435 (NRRL Y-11430). We hypothesized that other genetic backgrounds could harbor variants that affect yields of secreted proteins. We crossed CBS7435 with 2 other K . phaffii isolates and mapped quantitative trait loci (QTLs) for secretion of a heterologous protein, β-glucosidase, by sequencing individual segregant genomes. A major QTL mapped to a frameshift mutation in the mannosyltransferase gene HOC1 , which gives CBS7435 a weaker cell wall and higher protein secretion than the other isolates. Inactivation of HOC1 in the other isolates doubled β-glucosidase secretion. A second QTL mapped to an amino acid substitution in IRA1 that tripled β-glucosidase secretion in 1-week batch cultures but reduced cell viability, and its effects are specific to this heterologous protein. Our results demonstrate that QTL analysis is a powerful method for dissecting the basis of biotechnological traits in nonconventional yeasts, and a route to improving their industrial performance.