Redox robustness drives LPMO evolution

Area of Science:

• Biochemistry
• Enzymology
• Evolutionary Biology

Background:

• Lytic polysaccharide monooxygenases (LPMOs) are redox enzymes that utilize hydrogen peroxide (H2O2) in a peroxygenase reaction.
• These enzymes generate and utilize hydroxyl radicals, which can be damaging if not controlled.
• Understanding the evolution of LPMOs' catalytic abilities is crucial for comprehending metalloenzyme adaptation.

Purpose of the Study:

• To investigate the evolutionary steps behind the exceptional catalytic abilities of LPMOs.
• To elucidate how LPMOs evolved to control the generation and use of hydroxyl radicals.
• To understand the role of selective pressures in the evolution of metalloenzymes.

Main Methods:

• Ancestral sequence reconstruction and enzyme resurrection were employed.
• Real-time monitoring of copper reoxidation and amino acid radical formation was performed.
• Mutational studies of ancestral LPMOs were conducted.

Main Results:

• Evolutionary improvements were observed in avoiding futile H2O2 turnover and scavenging damaging radicals via a hole hopping pathway.
• Mutating ancestral LPMOs to adopt extant-like conformations in the hole hopping pathway enhanced redox robustness.
• Evolutionary pressures for generating potent oxidizing intermediates drive metalloenzyme evolution beyond the active site.

Conclusions:

• The evolution of LPMOs involved significant changes across the enzyme structure to manage reactive intermediates.
• The hole hopping pathway is critical for the redox robustness and catalytic efficiency of LPMOs.
• Metalloenzyme evolution is shaped by the functional demands of generating highly reactive species.