Proteomics Methodology Applied to the Analysis of Filamentous Fungi - New Trends for an Impressive Diverse Group of Organisms

A Review beyond the borders: Proteomics of microclonial black fungi and black yeasts

“…The fungus cell disruption is performed using different approaches by mechanical or enzymatic methodologies [14]. Since black fungi possess extraordinary rigid cell walls, the effective cell disruption is a crucial step for proteomic studies.…”

“…In this frame, the mechanical cell disruption can be combined with different homogenization buffers to enhance the disintegration of cell walls and inhibition of proteases activity [14]. Different concentrations of urea (7 -9 M) in combination with 2 M thiourea are used as a strong chaotropic to re-solubilize the proteins in filamentous fungi [21].…”

“…As a non-ionic detergent CHAPS is used at the concentrations of 1% -4%. It is recommendable to try different concentrations of CHAPS for the most desirable protein yield, which is to be optimized case to case [14]. DTT and ß-mercaptoethal were used as reducing agents in the homogenization buffer [24,25].…”

“…Enzymatic and chemical cell wall lysis was also used with less success for fungi samples [14]. Taken into consideration that black MCF are characterized by a more robust cell wall than filamentous fungi types, the solubilization of their proteins remain the main challenge for proteomic analyses.…”

“…The re-solubilization of protein pellet is significantly reduced after acidic precipitation, which affects the final protein yield. To avoid this problem the pellet must be dried very fast (1 -2 minutes) before re-solubilization [14]. The main obstacles in protein extraction from black fungi are the high complexity of fungi matrices containing melanins, pigments, polysaccharides, chitin and lipids.…”