Temporal and Spatial Resolution of a Protein Quake that Activates Hydrogen Tunneling in Soybean Lipoxygenase

Zaragoza, J. P.; Offenbacher, A. R.; Hu, S.; Gee, C. L.; Firestein, Z. M.; Minnetian, N.; Deng, Z.; Fan, F.; Iavarone, A. T.; Klinman, J. P.

doi:10.1101/2022.03.31.486235

bioRxiv · 10.1101/2022.03.31.486235

Temporal and Spatial Resolution of a Protein Quake that Activates Hydrogen Tunneling in Soybean Lipoxygenase

Zaragoza, J. P.·Offenbacher, A. R.·Hu, S.·Gee, C. L.·Firestein, Z. M.·Minnetian, N.·Deng, Z.·Fan, F.·Iavarone, A. T.·Klinman, J. P.

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Abstract

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 detect a radiating cone of aliphatic side chains that extends from the iron active site of SLO to the protein-solvent interface. Employing eight variants of SLO, nanosecond fluorescence Stokes shifts have been measured using a probe appended to the identified surface loop. We report a remarkable identity of the enthalpies of activation for the Stokes shifts decay rates and the millisecond C-H bond cleavage step that is restricted to side chain mutants within the identified thermal network. While the role of dynamics in enzyme function has been predominantly attributed to a distributed protein conformational landscape, these new data implicate a thermally initiated, cooperative protein quake as the source of the activation of SLO. These findings indicate a direct coupling of distal protein motions surrounding the exposed fluorescent probe to active site motions controlling catalysis.

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Zaragoza, J. P., Offenbacher, A. R., Hu, S., Gee, C. L., Firestein, Z. M., Minnetian, N., Deng, Z., Fan, F., Iavarone, A. T., Klinman, J. P.. 2022-04-01. Temporal and Spatial Resolution of a Protein Quake that Activates Hydrogen Tunneling in Soybean Lipoxygenase. https://doi.org/10.1101/2022.03.31.486235

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