Journal
JOURNAL OF STRUCTURAL BIOLOGY
Volume 205, Issue 1, Pages 22-29Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.jsb.2018.11.012
Keywords
Contrast transfer function; Focus gradient; Local focus refinement; Cryo-electron microscopy
Funding
- Discretionary Fund of the University of Michigan Life Sciences Institute
Ask authors/readers for more resources
Preferred particle orientation represents a recurring problem in single-particle cryogenic electron microcopy (cryo-EM). A specimen-independent approach through tilting has been attempted to increase particle orientation coverage, thus minimizing anisotropic three-dimensional (3D) reconstruction. However, focus gradient is a critical issue hindering tilt applications from being a general practice in single-particle cryo-EM. The present study describes a newly developed geometrically optimized approach, goCTF, to reliably determine the global focus gradient. A novel strategy of determining contrast transfer function (CTF) parameters from a sector of the signal preserved power spectrum is applied to increase reliability. Subsequently, per-particle based local focus refinement is conducted in an iterative manner to further improve the defocus accuracy. Novel diagnosis methods using a standard deviation defocus plot and goodness of fit heatmap have also been proposed to evaluate CTF fitting quality prior to 3D refmement. In a benchmark study, goC1P processed a published single particle cryo-EM dataset for influenza hemagglutinin trimer collected at a 40-degree specimen tilt. The resulting 3D reconstruction map was improved from 4.1 angstrom to 3.7 angstrom resolution. The goCTF program is built on the open source code of CTFFIND4, which adopts a consistent user interface for ease of use.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available