Atomic model of the F420-reducing [NiFe] hydrogenase by electron cryo-microscopy using a direct electron detector

Author:

Allegretti Matteo1,Mills Deryck J1,McMullan Greg2,Kühlbrandt Werner1,Vonck Janet1

Affiliation:

1. Department of Structural Biology, Max Planck Institute of Biophysics, Frankfurt, Germany

2. Medical Research Council Laboratory of Molecular Biology, Cambridge, United Kingdom

Abstract

The introduction of direct electron detectors with higher detective quantum efficiency and fast read-out marks the beginning of a new era in electron cryo-microscopy. Using the FEI Falcon II direct electron detector in video mode, we have reconstructed a map at 3.36 Å resolution of the 1.2 MDa F420-reducing hydrogenase (Frh) from methanogenic archaea from only 320,000 asymmetric units. Videos frames were aligned by a combination of image and particle alignment procedures to overcome the effects of beam-induced motion. The reconstructed density map shows all secondary structure as well as clear side chain densities for most residues. The full coordination of all cofactors in the electron transfer chain (a [NiFe] center, four [4Fe4S] clusters and an FAD) is clearly visible along with a well-defined substrate access channel. From the rigidity of the complex we conclude that catalysis is diffusion-limited and does not depend on protein flexibility or conformational changes.

Funder

Max Planck Society

UK Medical Research Council

Max-Planck-Gesellschaft

Medical Research Council

Publisher

eLife Sciences Publications, Ltd

Subject

General Immunology and Microbiology,General Biochemistry, Genetics and Molecular Biology,General Medicine,General Neuroscience

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