Secretome of the Coprophilous Fungus Doratomyces stemonitis C8, Isolated from Koala Feces

Abstract

Coprophilous fungi inhabit herbivore feces, secreting enzymes to degrade the most recalcitrant parts of plant biomass that have resisted the digestive process. Consequently, the secretomes of coprophilous fungi have high potential to contain novel and efficient plant cell wall degrading enzymes of biotechnological interest. We have used one-dimensional and two-dimensional gel electrophoresis, matrix-assisted laser desorption ionizationtime-of-flight tandem mass spectrometry (MALDI-TOF/TOF MS/MS), and quadrupole time-of-flight liquid chromatography-tandem mass spectrometry (Q-TOF LC-MS/MS) to identify proteins from the secretome of the coprophilous fungus Doratomyces stemonitis C8 (EU551185) isolated from koala feces. As the genome of D. stemonitis has not been sequenced, cross-species identification, de novo sequencing, and zymography formed an integral part of the analysis. A broad range of enzymes involved in the degradation of cellulose, hemicellulose, pectin, lignin, and protein were revealed, dominated by cellobiohydrolase of the glycosyl hydrolase family 7 and endo-1,4-␤-xylanase of the glycosyl hydrolase family 10. A high degree of specialization for pectin degradation in the D. stemonitis C8 secretome distinguishes it from the secretomes of some other saprophytic fungi, such as the industrially exploited T. reesei. In the first proteomic analysis of the secretome of a coprophilous fungus reported to date, the identified enzymes provide valuable insight into how coprophilous fungi subsist on herbivore feces, and these findings hold potential for increasing the efficiency of plant biomass degradation in industrial processes such as biofuel production in the future.Filamentous fungi exist in a broad range of habitats, fulfilling significant roles in a diversity of ecosystems. Integral to their survival is the ability to secrete enzymes to break down complex materials in the environment into small molecules that can be absorbed into the hyphae and used for nutrition (53). In the past decade, advances in protein identification techniques and genome sequencing have enabled detailed investigation of the secretomes of saprophytic (31, 42, 47, 50, 51), pathogenic (28, 35, 43), and symbiotic fungal species (29), revealing rich and diverse enzyme arrays. The fungal secretomes have been explored to find enzymes and enzyme combinations for various industrial applications, such as paper, textile, and food manufacture (7, 33) and economically and industrially sustainable hydrolysis of plant biomass to fermentable sugars for biofuel production (1, 13, 46, 51).Coprophilous fungi are a subgroup of saprophytic fungi that can inhabit feces, most commonly herbivore feces (53). As the waste product of the digestive process, herbivore feces are predominantly composed of the most recalcitrant and indigestible parts of the plant: the cell wall polymers cellulose, hemicellulose, and lignin (21). Therefore, the potential for the secretomes of coprophilous fungi to contain novel enzymes for efficient plant cell wall degradation is...

Fungal root endophytes ofQuercus robursubjected to flooding

“…; Peterson et al. ). The presence of coprophilous species in root tissues results from their evolution into plant‐associated fungi.…”

“…The Q. robur root endophytes recorded included two coprophilous species (Cephalotrichum stemonitis, Sordaria fimicola) and at least four entomopathogenic species (Clonostachys rosea, Isaria farinosa, I. fumosorosea, Metarhizium anisopliae). The coprophilous species may be transferred from faeces of herbivorous animals which are composed mostly of cellulose, hemicellulose and lignin, being degraded by a broad range of enzymes produced by fungi (Domsch et al 1980;Thormann et al 2002;Krug et al 2004;Peterson et al 2011). The presence of coprophilous species in root tissues results from their evolution into plant-associated fungi.…”

Analytical constraints for the analysis of human cell line secretomes by shotgun proteomics

“…The assessment of the full dynamic range of Saccharomyces cerevisiae and human urine proteomes exemplified the current approaches for quantifying proteins by selected reaction monitoring [9,10]. Recent shotgun proteomics reports revealed large catalogues of exoproteomes of several bacteria [11,12] and fungi [13] comprising a large list of items unsuspected from prior 2D-gel based studies. Regarding human proteomes, a large panel of secretomes have been described from myeloid cells [14], dendritic cells [15], stem cells [16], endothelial cells [17], astrocytes [18], and airway epithelial cells [19].…”

8 Degradation and Modification of Plant Biomass by Fungi

“…Growth of this fungus on plant-biomass-related substrates revealed secretion of enzymes related to cellulose and xylan degradation and to a lesser extent to mannan and pectin degradation, which matches the polysaccharides that are not or only partly degraded in the koala's gut (Peterson et al 2011). …”