Comparative secretome analyses of two Trichoderma reesei RUT-C30 and CL847 hypersecretory strains

ABSTRACT: BACKGROUND: Due to its capacity to produce large amounts of cellulases, Trichoderma reesei is increasingly been researched in various fields of white biotechnology, especially in biofuel production from lignocellulosic biomass. The commercial enzyme mixtures produced at industrial scales are not well characterized, and their proteinaceous components are poorly identified and quantified. The development of proteomic methods has made it possible to comprehensively overview the enzymes involved in lignocellulosic biomass degradation which are secreted under various environmental conditions. RESULTS: The protein composition of the secretome produced by industrial T. reesei (strain CL847) grown on a medium promoting the production of both cellulases and hemicellulases was explored using two-dimensional electrophoresis and MALDI-TOF or LC-MS/MS protein identification. A total of 22 protein species were identified. As expected, most of them are potentially involved in biomass degradation. The 2D map obtained was then used to compare the secretomes produced by CL847 and another efficient cellulolytic T. reesei strain, Rut-C30, the reference cellulase-overproducing strain using lactose as carbon source and inducer of cellulases. CONCLUSION: This study provides the most complete mapping of the proteins secreted by T. reesei to date. We report on the first use of proteomics to compare secretome composition between two cellulase-overproducing strains Rut-C30 and CL847 grown under similar conditions. Comparison of protein patterns in both strains highlighted many unexpected differences between cellulase cocktails. The results demonstrate that 2D electrophoresis is a promising tool for studying cellulase production profiles, whether for industrial characterization of an entire secretome or for a more fundamental study on cellulase expression at genome-wide scale

Quantitative determination of bovine kappa-casein macropeptide in dairy products by Liquid chromatography/Electrospray coupled to mass spectrometry (LC-ESI-MS) and Liquid chromatography/Electrospray coupled to tamdem mass spectrometry (LC-ESI/MS/MS)

κ-Casein macropeptide (CMP) is a polypeptide of 64 amino acid residues, derived from the C–terminal part of bovine κ-casein. It is a complex mixture of non-glycosylated and variously glycosylated forms. Using reversed phase—high pressure liquid chromatography (RP-HPLC) coupled to a mass spectrometer fitted with a electrospray ionisation source (ESI-MS) sensitive, reliable and specific methods were developed to enable unequivocal determination of aglyco-CMPA, aglyco-CMPB and total CMPAB. Several selective detection methods were proposed for their quantification. Aglyco-CMPA and aglyco-CMPB were quantified, respectively, from 13595+, 16984+, 22643+ and 13525+, 16904+, 22533+ m/z specific ions from multiple ion monitoring (MIM) mode. Within-day repeatability and between-day repeatability were 1% and 5%, respectively, with a standard deviation lower than 1%. Total CMPAB was quantified from peptide κ-CN (162–169) released from pepsin hydrolysis using UV, single ion monitoring (SIM) and multiple reaction monitoring (MRM) detection. A high degree of selectivity was obtained using MRM detection; the limit of quantification was 10 pmol within the standard curve range of 10–1000 pmol. Several commercial products were analysed and data obtained were in agreement with those indicated in the literature

Phosphopeptides interacting with colloidal calcium phosphate isolated by tryptic hydrolysis of bovine casein micelles

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Les peptidases des bactéries propioniques

International audienc

Peptides identified during emmental cheese ripening: origin and proteolytic systems involved

41 ref.International audienc

Modification of Bovine β-Lactoglobulin by Glycation in a Powdered State or in an Aqueous Solution: Effect on Association Behavior and Protein Conformation

The effect of glycation with lactose on the association behavior and conformational state of bovine β-lactoglobulin (β-LG) was studied, using size exclusion chromatography, polyacrylamide gel electrophoresis, proteolytic susceptibility, and binding of a fluorescent probe. Two modification treatments were used, i.e., aqueous solution glycation and dry-way glycation. The results showed that the latter treatment did not significantly alter the nativelike behavior of the protein while the former treatment led to important structural changes. These changes resulted in a specific denatured β-LG monomer, which covalently associated via the free thiol group. The homodimers thus formed and the expanded monomers underwent subsequent aggregation into a high molecular weight species, via noncovalent interactions. The association behavior of glycated β-LG is discussed with respect to the known multistep denaturation/aggregation process of nonmodified β-LG

Glycation de la beta-lactoglobuline par réaction de Maillard à l'aide de différents sucres : étude comparative des propriétés des polymères obtenus et des sites de substitution

International audienc

Peptides identified during emmental cheese ripening: origin and proteolytic systems involved

41 ref.International audienc

Les peptidases des bactéries propioniques

International audienc