Dose, exposure time, and resolution in Serial X-ray Crystallography

TL;DR

This paper analyzes the dose and exposure requirements in Serial X-ray Crystallography, demonstrating how resolution depends on flux, and predicts feasibility of high-resolution imaging with current and future X-ray sources.

Contribution

It introduces an analytical and simulation-based framework to evaluate X-ray dose, exposure time, and resolution in serial crystallography, highlighting the potential of fourth-generation sources for sub-nanometer resolution.

Findings

Resolution scales with the inverse fourth power of exposure time.

Sub-nanometer resolution is feasible with current synchrotrons using multiple beams.

Fourth-generation X-ray sources enable high-resolution imaging with shorter exposures.

Abstract

The resolution of X-ray diffraction microscopy is limited by the maximum dose that can be delivered prior to sample damage. In the proposed Serial Crystallography method, the damage problem is addressed by distributing the total dose over many identical hydrated macromolecules running continuously in a single-file train across a continuous X-ray beam, and resolution is then limited only by the available molecular and X-ray fluxes and molecular alignment. Orientation of the diffracting molecules is achieved by laser alignment. We evaluate the incident X-ray fluence (energy/area) required to obtain a given resolution from (1) an analytical model, giving the count rate at the maximum scattering angle for a model protein, (2) explicit simulation of diffraction patterns for a GroEL-GroES protein complex, and (3) the frequency cut off of the transfer function following iterative solution of the phase problem, and reconstruction of an electron density map in the projection approximation. These calculations include counting shot noise and multiple starts of the phasing algorithm. The results indicate counting time and the number of proteins needed within the beam at any instant for a given resolution and X-ray flux. We confirm an inverse fourth power dependence of exposure time on resolution, with important implications for all coherent X-ray imaging. We find that multiple single-file protein beams will be needed for sub-nanometer resolution on current third generation synchrotrons, but not on fourth generation designs, where reconstruction of secondary protein structure at a resolution of 0.7 nm should be possible with short exposures.