Quantitative proteomic analysis of secretome of microbial consortium during saw dust utilization Sunil S. Adav ⁎ , Anita Ravindran, Esther Sok Hwee Cheow, Siu Kwan Sze⁎⁎ School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, 637551, Singapore ARTICLE INFO ABSTRACT Article history: Received 27 May 2012 Accepted 13 August 2012 Available online 23 August 2012 Proteomics analysis of lignocellulolytic proteins by lignocellulosic biomass degrading microbes and compatible microbial consortium is a promising approach that offers a new means to enzyme discovery. The abundance of proteins in complex secretome by microbial communities would highlight key lignocellulolytic proteins for lignocellulosic biorefinery. In this study, lignocellulolytic enzymes of potent lignin degrading basidiomycota and effective cellulolytic ascomycota fungal strains, and their co-cultures were analyzed using high throughput isobaric tag for relative and absolute quantitation (iTRAQ) technique using liquid chromatography tandem mass spectrometry. Protein abundances in the iTRAQ- multiplexed samples were determined by integrating relative quantitation and exponentially modified protein abundance index (emPAI). The functional classification of the secretory proteins by individual culture and co-culture demonstrated 36.77% cellulolytic proteins, 13.06% hemicellulases, 14.09% ligninolytic proteins, 19.59% proteolytic enzymes. 7.22% hypothetical proteins and 6.87% cell morphogenesis proteins. The abundance of the proteins by individual cultures and co-cultured fungal consortium revealed that co- culturing of Phanerochaete chrysosporium with Trichoderma reesei QM6a and Trichoderma reesei Rut C30 induced the production of cellulolytic proteins and stimulated expression of hemicellulolytic enzymes. The hierarchical clustering of proteins in secretome of fungal strains and their co-cultures elucidated differential expressions of lignocellulolytic proteins by the microbial consortium. © 2012 Elsevier B.V. All rights reserved. Keywords: Phanerochaete chrysosporium Trichoderma reesei Secretome iTRAQ Lignocellulosic biomass Microbial consortium 1. Introduction Lignocellulosic biomass is an important and abundant renew- able nature's carbon basin for the biorefinery industry. Given its sustainability and environmental-friendliness, lignocellu- losic biomass has been recognized as the potential alternative to the current petroleum industry. The plant biomass includ- ing agricultural residues and forest wastes consists of cellulose, hemicellulose, pectin and recalcitrant lignin. The carbohydrate constituent of biomass can be hydrolyzed to monomeric sugars and the resulting hydrolysate may serve as versatile substrates for fermentation or biocatalytic processes in the production of biofuels, biopolymers and chemicals. To improve lignocellulosic biomass hydrolysis, different biomass pretreatment methods, microbial enzymatic processes and genetic engineering of plant to reduce lignin content were documented [1–3]. Pretreatment processes are expensive and generate various compounds such as organic acid, furfurals and phenolic compounds that interfere in microbial activities [4]. On the contrary, several microorganisms from both fungal and bacterial domain degrade lignocellulosic biomass, with significant contribution by fungi, hence fungal extracellular enzymes as well as metabolic processes are under investiga- tion [5–12]. JOURNAL OF PROTEOMICS 75 (2012) 5590 – 5603 ⁎ Corresponding author. Tel.: +65 6316 2852; fax: +65 6791 3856. ⁎⁎ Corresponding author. Tel.: +65 6514 1006; fax: +65 6791 3856. E-mail addresses: ssadav@ntu.edu.sg (S.S. Adav), sksze@ntu.edu.sg (S.K. Sze). 1874-3919/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jprot.2012.08.011 Available online at www.sciencedirect.com www.elsevier.com/locate/jprot