Genome-scale analysis of the high-efficient protein secretion system of Aspergillus oryzae

Abstract

BackgroundThe koji mold, Aspergillus oryzae is widely used for the production of industrial enzymes due to its particularly high protein secretion capacity and ability to perform post-translational modifications. However, systemic analysis of its secretion system is lacking, generally due to the poorly annotated proteome.ResultsHere we defined a functional protein secretory component list of A. oryzae using a previously reported secretory model of S. cerevisiae as scaffold. Additional secretory components were obtained by blast search with the functional components reported in other closely related fungal species such as Aspergillus nidulans and Aspergillus niger. To evaluate the defined component list, we performed transcriptome analysis on three α-amylase over-producing strains with varying levels of secretion capacities. Specifically, secretory components involved in the ER-associated processes (including components involved in the regulation of transport between ER and Golgi) were significantly up-regulated, with many of them never been identified for A. oryzae before. Furthermore, we defined a complete list of the putative A. oryzae secretome and monitored how it was affected by overproducing amylase.ConclusionIn combination with the transcriptome data, the most complete secretory component list and the putative secretome, we improved the systemic understanding of the secretory machinery of A. oryzae in response to high levels of protein secretion. The roles of many newly predicted secretory components were experimentally validated and the enriched component list provides a better platform for driving more mechanistic studies of the protein secretory pathway in this industrially important fungus.

Comparative RNA-seq based transcriptomic analysis of Aspergillus nidulans recombinant strains overproducing heterologous glycoside hydrolases

“…These subcategories were also reported in other Aspergillus species overproducing recombinant proteins 27,38,39 . The overexpression of glucoamylase (GlaA) in A. niger resulted in the activation of "translocation", "protein glycosylation", "vesicle transport" and "ion homeostasis" processes 38 .…”

“…The production of recombinant proteins in filamentous fungi by using strong promoters activates some regulatory genes in pathways related to protein quality control and UPR [26][27][28] . The UPR represents an adaptive response that aims to restore cellular homeostasis triggered by ER stress 29 .…”

“…The production of heterologous proteins is not only impaired by low expression of heterologous genes, but it can also be hampered by problems in post-translational processing in the secretory pathway 50 . Many studies have attempted to comprehend the superior secretion capacity of Aspergillus, primarily at the transcriptomic level 6,16,25,27,38 . These studies identified important genes in different stages of protein production such as translocation, folding, cargo transport and exocytosis 51 .…”

Systematic Optimization of Protein Secretory Pathways in Saccharomyces cerevisiae to Increase Expression of Hepatitis B Small Antigen

“…Another transcriptome analysis on three α-amylase over-producing Aspergillus oryzae strains has also identified a complete list of the putative secretome and confirmed its effect on the overproduction of amylase (Liu et al, 2014). Because the protein secretory pathways are conserved in eukaryotic organisms, we hypothesized in this study that genes involved in protein secretory pathways could be rate limiting for protein expression in S. cerevisiae .…”

Recombinant expression and purification of an Oxysterol Binding Protein fromAspergillus oryzae3.042

“…[2]. The genome of A. oryzae has been sequenced in 2005 [3], which has facilitated research efforts to understand its basic biology and develop better industrial strains of this significant fungus [4] .…”