Deficiency of biodegradable plastic-degrading enzyme production in a gene-deletion mutant of phyllosphere yeast, Pseudozyma antarctica defective in mannosylerythritol lipid biosynthesis

Abstract

Abstract The basidiomycetous yeast Pseudozyma antarctica (currently designated Moesziomyces antarcticus) produces extracellular enzymes and glycolipids, including mannosylerythritol lipids (MELs), which are biosurfactants. Strain GB-4(0) of this species was previously isolated from rice husks and produces biodegradable plastic-degrading enzyme (Pseudozyma antarctica esterase; PaE). In this study, we generated a MEL biosynthesis-deficient strain (∆PaEMT1) by deleting the gene PaEMT1, which is essential to MEL biosynthesis in strain GB-4(0). The resulting ∆PaEMT1 strain showed deficient PaE activity, and the corresponding signal was hardly detected in its culture supernatant through western blotting analysis using rabbit anti-PaE serum. On the other hand, the relative expression of the gene PaCLE1, encoding PaE, was identical between GB-4(0) and ∆PaEMT1 based on quantitative real-time PCR. When strain ∆PaEMT1 was grown in culture media supplemented with various surfactants, i.e., Tween20, BRIJ35 and TritonX-100, and MELs, PaE activity and secretion recovered. We also attempted to detect intracellular PaE using cell-free extract, but observed no signal in the soluble or insoluble fractions of ∆PaEMT1. This result suggested that the PaCLE1 gene was not translated to PaE, or that expressed PaE was degraded immediately in ∆PaEMT1. Based on these results, MEL biosynthesis is an important contributor to PaE production.

Keywords

• Pseudozyma antarctica
• Esterase
• Biodegradable plastic
• Mannosylerythritol lipid
• Glycosyltransferase
• Gene deletion