Florian Csarman, M.Sc.
Current position: Ph.D. Candidate
Project Funding: OXIDISE
Home University: BOKU
Duration of Stay: 10/2017-09/2020
The aim of this study is the investigation of the lignocellulolytic machinery of basidiomycetous brown-rot fungi, due to their important role in the carbon cycle of forests as efficient degraders of plant biomass.
The first phase of this PhD-project focusses on the heterologous expression of secreted oxidoreductases and glycoside hydrolases of Fomitopsis pinicola in Trichoderma reesei. Heterologous enzymes are characterized biochemically to verify their natural activities and homogenous preparations of the enzymes are further investigated concerning their interaction kinetics by steady-state and stopped-flow methods. Electrochemical methods will be applied to study all major oxidoreductases concerning electron transfer, regeneration of redox species and substrate cascading.
For the further elucidation of the interactions between enzymes and their natural substrates, surface plasmon resonance (SPR) is the method of choice. For this purpose, sensor chips modified with different lignin, cellulose and hemicellulose preparations are produced in-house to determine binding constants and kinetics. The spatially resolved detection of enzymes and reaction products at high resolution at the surface of their natural substrates is achieved using specific biosensors. Scanning electrochemical microscopy (SECM) using miniaturized electrodes is a potent method for the investigation of the distribution of enzymes and intermediates of the degradation process.
In the final phase of the project the previously developed techniques using purified enzyme preparations should be adapted and further modified to study the enzyme distribution directly at the fungal hyphae in vivo.