Temporal and spatial resolution of distal protein motions that activate hydrogen tunneling in soybean lipoxygenase.
Jan Paulo T ZaragozaAdam R OffenbacherShenshen HuChristine L GeeZachary M FiresteinNatalie MinnetianZhenyu DengFlora FanAnthony T IavaroneJudith P KlinmanPublished in: Proceedings of the National Academy of Sciences of the United States of America (2023)
The enzyme soybean lipoxygenase (SLO) provides a prototype for deep tunneling mechanisms in hydrogen transfer catalysis. This work combines room temperature X-ray studies with extended hydrogen-deuterium exchange experiments to define a catalytically-linked, radiating cone of aliphatic side chains that connects an active site iron center of SLO to the protein-solvent interface. Employing eight variants of SLO that have been appended with a fluorescent probe at the identified surface loop, nanosecond fluorescence Stokes shifts have been measured. We report a remarkable identity of the energies of activation ( E a ) for the Stokes shifts decay rates and the millisecond C-H bond cleavage step that is restricted to side chain mutants within an identified thermal network. These findings implicate a direct coupling of distal protein motions surrounding the exposed fluorescent probe to active site motions controlling catalysis. While the role of dynamics in enzyme function has been predominantly attributed to a distributed protein conformational landscape, the presented data implicate a thermally initiated, cooperative protein reorganization that occurs on a timescale faster than nanosecond and represents the enthalpic barrier to the reaction of SLO.
Keyphrases
- fluorescent probe
- living cells
- room temperature
- protein protein
- amino acid
- single molecule
- binding protein
- minimally invasive
- ionic liquid
- magnetic resonance imaging
- computed tomography
- high resolution
- molecular dynamics
- gene expression
- magnetic resonance
- big data
- dna methylation
- electronic health record
- artificial intelligence
- dna binding
- machine learning
- quantum dots