Abstract
The computational design of a symmetric protein homo-oligomer that binds a symmetry-matched small molecule larger than a metal ion has not yet been achieved. We used de novo protein design to create a homo-trimeric protein that binds the C3 symmetric small molecule drug amantadine with each protein monomer making identical interactions with each face of the small molecule. Solution NMR data show that the protein has regular three-fold symmetry and undergoes localized structural changes upon ligand binding. A high-resolution X-ray structure reveals a close overall match to the design model with the exception of water molecules in the amantadine binding site not included in the Rosetta design calculations, and a neutron structure provides experimental validation of the computationally designed hydrogen-bond networks. Exploration of approaches to generate a small molecule inducible homo-trimerization system based on the design highlight challenges that must be overcome to computationally design such systems.
| Original language | English |
|---|---|
| Article number | e47839 |
| Journal | eLife |
| Volume | 8 |
| DOIs | |
| State | Published - Dec 2019 |
Funding
We thank Dave Roberts (DePauw University) and Norma Dukes (SBC) for assistance with X-ray diffraction data collection and data processing. Results shown in this report are derived from work performed at Argonne National Laboratory, Structural Biology Center (SBC) at the Advanced Photon Source. SBC-CAT is operated by UChicago Argonne, LLC, for the US Department of Energy, Office of Biological and Environmental Research under contract DE-AC02-06CH11357. Neutron diffraction data were collected at the High Flux Isotope Reactor, a DOE Office of Science User Facility operated by the Oak Ridge National Laboratory. NMR data acquisition was supported through the Office of the Director, NIH, under High End Instrumentation (HIE) Grant S10OD018455, which funded the 800 MHz NMR spectrometer at UCSC. JP is supported by the Washington Research Foundation Innovation Postdoctoral Fellowship. ACM and N.GS are supported by an R35 Outstanding Investigator Award through NIGMS(1R35GM125034-01). SEB was supported by the Burroughs Wellcome Fund Career Award at the Scientific Interface. This work was also supported by HHMI. The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies.
| Funders | Funder number |
|---|---|
| DePauw University | |
| Office of Biological and Environmental Research | |
| US Department of Energy | |
| National Institutes of Health | S10OD018455 |
| Howard Hughes Medical Institute | |
| U.S. Department of Energy | |
| National Institute of General Medical Sciences | R35GM125034 |
| Office of the Director | |
| Burroughs Wellcome Fund | |
| Washington Research Foundation | |
| Interface | |
| Office of Science | |
| Biological and Environmental Research | DE-AC02-06CH11357 |
| Argonne National Laboratory | |
| Oak Ridge National Laboratory | |
| University of California, Santa Cruz | |
| University of Chicago | |
| Anacostia Community Museum | |
| Society for Experimental Biology |