Format

Send to

Choose Destination
Nature. 2018 Sep;561(7724):485-491. doi: 10.1038/s41586-018-0509-0. Epub 2018 Sep 12.

De novo design of a fluorescence-activating β-barrel.

Author information

1
Department of Biochemistry, University of Washington, Seattle, WA, USA.
2
Institute for Protein Design, University of Washington, Seattle, WA, USA.
3
Division of Basic Science, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
4
Crown Bioscience, Taicang, China.
5
Department of Chemistry, University of Washington, Seattle, WA, USA.
6
Molecular Biophysics and Integrated Bioimaging, Berkeley Center for Structural Biology, Lawrence Berkeley National Laboratory, Berkeley, CA, USA.
7
John Harvard Distinguished Science Fellowship Program, Harvard University, Cambridge, MA, USA.
8
Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology, Barcelona, Spain.
9
Department of Bioengineering, Stanford University, Stanford, CA, USA.
10
Department of Biochemistry, University of Washington, Seattle, WA, USA. dabaker@uw.edu.
11
Institute for Protein Design, University of Washington, Seattle, WA, USA. dabaker@uw.edu.
12
Howard Hughes Medical Institute, University of Washington, Seattle, WA, USA. dabaker@uw.edu.

Abstract

The regular arrangements of β-strands around a central axis in β-barrels and of α-helices in coiled coils contrast with the irregular tertiary structures of most globular proteins, and have fascinated structural biologists since they were first discovered. Simple parametric models have been used to design a wide range of α-helical coiled-coil structures, but to date there has been no success with β-barrels. Here we show that accurate de novo design of β-barrels requires considerable symmetry-breaking to achieve continuous hydrogen-bond connectivity and eliminate backbone strain. We then build ensembles of β-barrel backbone models with cavity shapes that match the fluorogenic compound DFHBI, and use a hierarchical grid-based search method to simultaneously optimize the rigid-body placement of DFHBI in these cavities and the identities of the surrounding amino acids to achieve high shape and chemical complementarity. The designs have high structural accuracy and bind and fluorescently activate DFHBI in vitro and in Escherichia coli, yeast and mammalian cells. This de novo design of small-molecule binding activity, using backbones custom-built to bind the ligand, should enable the design of increasingly sophisticated ligand-binding proteins, sensors and catalysts that are not limited by the backbone geometries available in known protein structures.

PMID:
30209393
PMCID:
PMC6275156
[Available on 2019-03-12]
DOI:
10.1038/s41586-018-0509-0

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center