Interdomain electron transfer in cellobiose dehydrogenase is governed by surface electrostatics

Area of Science:

• Biochemistry
• Structural Biology
• Biotechnology

Background:

• Cellobiose dehydrogenase (CDH) is a fungal oxidoreductase crucial for cellulose degradation.
• CDH facilitates electron transfer to lytic polysaccharide monooxygenases.
• The pH-dependent mechanism of interdomain electron transfer in CDH remained experimentally unproven.

Purpose of the Study:

• To elucidate the structural basis of domain interaction in CDH.
• To investigate the pH-dependent regulation of interdomain electron transfer.
• To understand the role of electrostatics in CDH functional states.

Main Methods:

• Hydrogen/deuterium exchange mass spectrometry (HDX-MS) with an optimized proteolytic setup.
• Native mass spectrometry coupled with ion mobility.
• Computational electrostatics calculations.

Main Results:

• HDX-MS showed pH-dependent alterations in solvent accessibility and hydrogen bonding at the CDH interdomain interface.
• Electrostatics calculations and ion mobility revealed increased interdomain electrostatic repulsion at neutral pH due to protonation-induced charge neutralization.
• Extensive O-glycosylation in the linker region and the precise papain cleavage site were identified.

Conclusions:

• CDH transitions between inactive (open) and active (closed) states are governed by surface electrostatics at the domain interface.
• Interdomain electrostatic repulsion is the primary modulator of CDH function.
• This study provides experimental evidence for charge repulsion's role in CDH interdomain electron transfer, relevant for biosensors and biofuel cells.