Enzyme intermediates captured “on the fly” by mix-and-inject serial crystallography
Posted on 2018-05-31 - 05:00
Abstract Background Ever since the first atomic structure of an enzyme was solved, the discovery of the mechanism and dynamics of reactions catalyzed by biomolecules has been the key goal for the understanding of the molecular processes that drive life on earth. Despite a large number of successful methods for trapping reaction intermediates, the direct observation of an ongoing reaction has been possible only in rare and exceptional cases. Results Here, we demonstrate a general method for capturing enzyme catalysis “in action” by mix-and-inject serial crystallography (MISC). Specifically, we follow the catalytic reaction of the Mycobacterium tuberculosis β-lactamase with the third-generation antibiotic ceftriaxone by time-resolved serial femtosecond crystallography. The results reveal, in near atomic detail, antibiotic cleavage and inactivation from 30 ms to 2 s. Conclusions MISC is a versatile and generally applicable method to investigate reactions of biological macromolecules, some of which are of immense biological significance and might be, in addition, important targets for structure-based drug design. With megahertz X-ray pulse rates expected at the Linac Coherent Light Source II and the European X-ray free-electron laser, multiple, finely spaced time delays can be collected rapidly, allowing a comprehensive description of biomolecular reactions in terms of structure and kinetics from the same set of X-ray data.
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Olmos, Jose; Pandey, Suraj; Martin-Garcia, Jose; Calvey, George; Katz, Andrea; Knoska, Juraj; et al. (2018). Enzyme intermediates captured “on the fly” by mix-and-inject serial crystallography. figshare. Collection. https://doi.org/10.6084/m9.figshare.c.4117712.v1
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AUTHORS (50)
JO
Jose Olmos
SP
Suraj Pandey
JM
Jose Martin-Garcia
GC
George Calvey
AK
Andrea Katz
JK
Juraj Knoska
CK
Christopher Kupitz
MH
Mark Hunter
ML
Mengning Liang
DO
Dominik Oberthuer
OY
Oleksandr Yefanov
MW
Max Wiedorn
MH
Michael Heyman
MH
Mark Holl
KP
Kanupriya Pande
AB
Anton Barty
MM
Mitchell Miller
SS
Stephan Stern
SR
Shatabdi Roy-Chowdhury
JC
Jesse Coe
KEYWORDS
biomolecular reactionscrystallography Abstract Backgroundstructure-based drug designEnzyme intermediatesLinac Coherent Light Source IImegahertz X-ray pulse ratesdrive lifeMycobacterium tuberculosis β- lactamasefemtosecond crystallographymethod30 msConclusions MISCreaction intermediatesEuropean X-ray free-electron laserantibiotic cleavageantibiotic ceftriaxoneX-ray datatime delays