Determining the Mechanism of DJ-1 Using Mix-and-Inject Synchrotron Serial Crystallography
Category
Published on
Type
journal-article
Author
Cole Dolamore and Kara Zielinski and Kevin Dalton and Stephen Meisburger and John Termini and Nathan Smith and Robert Henning and Vukica Srajer and Doeke Hekstra and Lois Pollack and Mark Wilson
Citation
Dolamore, C., Zielinski, K., Dalton, K., Meisburger, S., Termini, J., Smith, N., Henning, R., Srajer, V., Hekstra, D., Pollack, L., & Wilson, M. (2025). Determining the Mechanism of DJ-1 Using Mix-and-Inject Synchrotron Serial Crystallography. Structural Dynamics, 12(2_Supplement), A69–A69. https://doi.org/10.1063/4.0000378
Abstract
Determining the molecular basis of enzyme catalytic activity is a central challenge in biochemistry. Time-resolved serial X-ray crystallography using X-ray free electron (XFELs) and synchrotron sources allows enzyme reactions to be observed in real time in a crystalline environment (1-3). We used pink beam mix-and-inject serial crystallography (MISC) at the Advanced Photon Source (BioCARS 14-ID) to follow the conversion of methylglyoxal to L-lactate by human DJ-1 using serial Laue diffraction. The high flux of the pink X-ray beam permitted microsecond exposure times, in contrast to millisecond exposure times typical at monochromatic beamlines. DJ-1's enzymatic activity has been controversial, with proposals that it functions either as a direct glyoxalase or as a deglycase acting only on glycated macromolecular substrates. Using custom-built injectors, we collected a time series of datasets and observed the formation of covalent catalytic intermediates in DJ-1 microcrystals after mixing with methylglyoxal (4). Our results establish that DJ-1 acts directly on methylglyoxal and demonstrate the feasibility of performing time-resolved MISC experiments at synchrotron beamlines.
