4WHA

Lipoxygenase-1 (soybean) L546A/L754A mutant

Summary for 4WHA

• Entry DOI: 10.2210/pdb4wha/pdb
• Related: 3PZW
• Descriptor: Seed linoleate 13S-lipoxygenase-1, FE (III) ION, 1,2-ETHANEDIOL, ... (5 entities in total)
• Functional Keywords: lipoxygenase, tunneling, c-h activation, oxidoreductase
• Biological source: Glycine max (Soybean)
• Total number of polymer chains: 1
• Total formula weight: 95400.67

Authors

Scouras, A.D.,Carr, C.A.M.,Hu, S.,Klinman, J.P. (deposition date: 2014-09-21, release date: 2014-11-12, Last modification date: 2023-09-27)

Primary citation

Hu, S.,Sharma, S.C.,Scouras, A.D.,Soudackov, A.V.,Carr, C.A.,Hammes-Schiffer, S.,Alber, T.,Klinman, J.P.
Extremely elevated room-temperature kinetic isotope effects quantify the critical role of barrier width in enzymatic C-H activation.
J.Am.Chem.Soc., 136:8157-8160, 2014

PubMed Abstract: The enzyme soybean lipoxygenase (SLO) has served as a prototype for hydrogen-tunneling reactions, as a result of its unusual kinetic isotope effects (KIEs) and their temperature dependencies. Using a synergy of kinetic, structural, and theoretical studies, we show how the interplay between donor-acceptor distance and active-site flexibility leads to catalytic behavior previously predicted by quantum tunneling theory. Modification of the size of two hydrophobic residues by site-specific mutagenesis in SLO reduces the reaction rate 10(4)-fold and is accompanied by an enormous and unprecedented room-temperature KIE. Fitting of the kinetic data to a non-adiabatic model implicates an expansion of the active site that cannot be compensated by donor-acceptor distance sampling. A 1.7 Å resolution X-ray structure of the double mutant further indicates an unaltered backbone conformation, almost identical side-chain conformations, and a significantly enlarged active-site cavity. These findings show the compelling property of room-temperature hydrogen tunneling within a biological context and demonstrate the very high sensitivity of such tunneling to barrier width.

PubMed: 24884374
DOI: 10.1021/ja502726s

Experimental method

X-RAY DIFFRACTION (1.7 Å)