9BJS

X-ray crystal structure of Y62W Thermothelomyces thermophilus polysaccharide monooxygenase 9E

Summary for 9BJS

• Entry DOI: 10.2210/pdb9bjs/pdb
• Related: 9BJQ 9BJR
• Descriptor: Glycoside hydrolase family 61 protein, COPPER (II) ION, 1,2-ETHANEDIOL, ... (6 entities in total)
• Functional Keywords: polysaccharide monooxygenase, oxidoreductase
• Biological source: Thermothelomyces thermophilus
• Total number of polymer chains: 2
• Total formula weight: 49141.15

Authors

Thomas, W.C.,Sayler, R.I.,Marletta, M.A. (deposition date: 2024-04-25, release date: 2024-12-18, Last modification date: 2025-01-22)

Primary citation

Sayler, R.I.,Thomas, W.C.,Rose, A.J.,Marletta, M.A.
Electron transfer in polysaccharide monooxygenase catalysis.
Proc.Natl.Acad.Sci.USA, 122:e2411229121-e2411229121, 2025

PubMed Abstract: Polysaccharide monooxygenase (PMO) catalysis involves the chemically difficult hydroxylation of unactivated C-H bonds in carbohydrates. The reaction requires reducing equivalents and will utilize either oxygen or hydrogen peroxide as a cosubstrate. Two key mechanistic questions are addressed here: 1) How does the enzyme regulate the timely and tightly controlled electron delivery to the mononuclear copper active site, especially when bound substrate occludes the active site? and 2) How does this electron delivery differ when utilizing oxygen or hydrogen peroxide as a cosubstrate? Using a computational approach, potential paths of electron transfer (ET) to the active site copper ion were identified in a representative AA9 family PMO from (PMO9E). When Y62, a buried residue 12 Å from the active site, is mutated to F, lower activity is observed with O. However, a WT-level activity is observed with HO as a cosubstrate indicating an important role in ET for O activation. To better understand the structural effects of mutations to Y62 and axial copper ligand Y168, crystal structures were solved of the wild type PMO9E and the variants Y62W, Y62F, and Y168F. A bioinformatic analysis revealed that position 62 is conserved as either Y or W in the AA9 family. The PMO9E Y62W variant has restored activity with O. Overall, the use of redox-active residues to supply electrons for the reaction with O appears to be widespread in the AA9 family. Furthermore, the results provide a molecular framework to understand catalysis with O versus HO.

PubMed: 39793048
DOI: 10.1073/pnas.2411229121

Experimental method

X-RAY DIFFRACTION (2 Å)