Research Article

Proton uptake mechanism in bacteriorhodopsin captured by serial synchrotron crystallography

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Science  05 Jul 2019:
Vol. 365, Issue 6448, pp. 61-65
DOI: 10.1126/science.aaw8634

Refilling the proton pump

Proteins are dynamic. Rearrangements of side chains, secondary structure, and entire domains gate functional transitions on time scales ranging from picoseconds to milliseconds. Weinert et al. used time-resolved serial crystallography to study large conformational changes in the proton pump bacteriorhodopsin that allow for redistribution of protons during the pumping cycle. They adapted methods used for x-ray free electron lasers to synchrotron x-ray sources. Large loop movements and a chain of water molecules were central to regenerating the starting state of bacteriorhodopsin.

Science, this issue p. 61


Conformational dynamics are essential for proteins to function. We adapted time-resolved serial crystallography developed at x-ray lasers to visualize protein motions using synchrotrons. We recorded the structural changes in the light-driven proton-pump bacteriorhodopsin over 200 milliseconds in time. The snapshot from the first 5 milliseconds after photoactivation shows structural changes associated with proton release at a quality comparable to that of previous x-ray laser experiments. From 10 to 15 milliseconds onwards, we observe large additional structural rearrangements up to 9 angstroms on the cytoplasmic side. Rotation of leucine-93 and phenylalanine-219 opens a hydrophobic barrier, leading to the formation of a water chain connecting the intracellular aspartic acid–96 with the retinal Schiff base. The formation of this proton wire recharges the membrane pump with a proton for the next cycle.

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