Published in
Nature Research, Scientific Reports, 1(5), 2015
DOI: 10.1038/srep14290
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Tools
Cryo-EM and the elucidation of new macromolecular structures: Random Conical Tilt revisited
,
Martín Alcorlo,
J. M. de la Rosa-Trevín,
José Miguel De La Rosa-Trevín,
Roberto Melero ,
I. Foche,
A. Zaldívar-Peraza,
L. del Cano,
Javier Vargas,
Vahid Abrishami,
Joaquín Otón ,
Roberto Marabini ,
José María Carazo
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Abstract
AbstractCryo-Electron Microscopy (cryo-EM) of macromolecular complexes is a fundamental structural biology technique which is expanding at a very fast pace. Key to its success in elucidating the three-dimensional structure of a macromolecular complex, especially of small and non-symmetric ones, is the ability to start from a low resolution map, which is subsequently refined with the actual images collected at the microscope. There are several methods to produce this first structure. Among them, Random Conical Tilt (RCT) plays a prominent role due to its unbiased nature (it can create an initial model based on experimental measurements). In this article, we revise the fundamental mathematical expressions supporting RCT, providing new expressions handling all key geometrical parameters without the need of intermediate operations, leading to improved automation and overall reliability, essential for the success of cryo-EM when analyzing new complexes. We show that the here proposed RCT workflow based on the new formulation performs very well in practical cases, requiring very few image pairs (as low as 13 image pairs in one of our examples) to obtain relevant 3D maps.