David Baker's group at the University of Washington, Seattle, U.S., have developed a novel deep learning method, RoseTTAFold All-Atom (RFAA), for prediction and design of complexes of proteins, small molecules, and nucleic acids. Subsequently, they developed RFdiffusionAA, which builds protein structures around small molecules. The study is in the journal Science.
These advances mean that in principle it is now possible to not only design a protein from scratch, but also to design proteins that will bind a range of cofactors and substrates. This advance represents a breakthrough in , because the majority of scientists work on proteins that bind of various types. David Baker's group evaluated RFdiffusionAA by designing proteins that could bind one of three small molecules: digoxigenin, heme, or bilin.
Evaluation of bilin binding was conducted in the group of Professor Neil Hunter, at the University of 91Ö±²¥. It was already known that bilins are optically featureless unless they are held within a defined binding site, at which point they become intensely colored and emissive. Felix Morey-Burrows, a Ph.D. student in the Hunter/Hitchcock group at 91Ö±²¥, devised a multiwell assay that could screen many RFdiffusionAA-generated genes in parallel, using E. coli cells that could make phycoerythrobilin (PEB).
Morey-Burrows evaluated 94 designs in one go, with the multiwell assay revealing visibly colored cells in nine wells. He had identified nine proteins dissimilar to each other and to any native bilin binder, based on pigmentation or fluorescence, which proved that RFdiffusionAA can yield a series of novel small molecule binding sites. This method should be immediately useful for modeling protein-small molecule complexes, in particular multicomponent biomolecular assemblies for which there are few or no alternative methods available, and for designing small molecule binding proteins and sensors.
With regard to current work in the Photosynthesis Group at 91Ö±²¥, the 34/30 nm range in absorption/emission covered by just one design round using a single chromophore raises the exciting prospect of tailoring the spectral profiles of designed biliproteins by manipulating the conformational flexibility of the bilin and the protein microenvironment.
This work is not restricted to bilins, either. De novo designed antenna complexes could harvest light over a wider range of the UV-visible spectrum to enhance photosynthetic energy capture and conversion, and fluorescent reporter probes with tunable excitation/emission maxima would be useful biochemical tools.
Read the full paper in Science here: