9TI1

Crystal structure of the zinc-containing Phosphotriesterase dPTE2-H55

Summary for 9TI1

• Entry DOI: 10.2210/pdb9ti1/pdb
• Descriptor: Parathion hydrolase, ZINC ION, 1,2-ETHANEDIOL, ... (4 entities in total)
• Functional Keywords: phosphotriesterase, zinc-dependent, hydrolase
• Biological source: Brevundimonas diminuta
• Total number of polymer chains: 2
• Total formula weight: 74797.55

Authors

Manser, B.P.,Deliz Liang, A. (deposition date: 2025-12-04, release date: 2026-02-25, Last modification date: 2026-03-11)

Primary citation

Manser, B.P.,Deliz Liang, A.
Catalytic p K a Attenuation in a Hydrolytic Metalloenzyme by Genetic Code Expansion.
Biochemistry, 65:559-570, 2026

PubMed Abstract: Hydrolytic metalloenzymes employ Lewis-acidic metal cofactors to activate water molecules, generating nucleophilic hydroxide species that facilitate catalysis. Their catalytic efficiency across a wide pH range is often governed by the protonation state of the metal-bound water, reflected in p values typically between 6.8 and 9. Modulating this parameter is key to expanding enzymatic activity for improved activity at neutral to acidic pH. Herein, we apply genetic code expansion to mutate the primary metal-coordination sphere of a model metallohydrolase: the dizinc phosphotriesterase from . Substitution of the most catalytically indispensable coordinating histidine residue (H55) to -methyl-l-histidine (πMH) resulted in substantial enzyme yields, efficient metal coordination for either Zn or Co, and up to 5-fold improved tolerance to acidic conditions. Detailed mechanistic analysis revealed a systematic decrease in catalytic p and attenuation of several catalytic rate constants. These results add to the growing body of evidence demonstrating the power of ncAA-based engineering for refined tuning of enzyme properties.

PubMed: 41705832
DOI: 10.1021/acs.biochem.5c00768

Experimental method

X-RAY DIFFRACTION (1.55 Å)