A Breakthrough in Protein Design

A computational method to design and create proteins that bind to a specific target could give rise to new drugs.

This is from Codon, my weekly newsletter. Subscribe for free.

Hello. A computational method to design and create proteins that bind to a specific target could give rise to new drugs. Engineered bacteria, injected into mice, target and destroy tumors using pulses of ultrasound. An improved prime editing protein swaps nucleotides in plants with high efficiency. And a CRISPR tool could help unearth hidden metabolites in the bacterial world teeming beneath our feet.

Protein Design, Backwards

The human genome encodes about 20,000 proteins, each of which is a three-dimensional jigsaw puzzle with pockets, folds, crevices, cracks. The ability to look at a protein's three-dimensional structure, and then reverse engineer a new protein that can bind to that target and block its function — called a binder — would be truly transformative for creating new drugs. A study, published last week in Nature, offers a computational method to do precisely that.

With some high-end computers and clever thinking, researchers in David Baker's lab at the Institute for Protein Design in Seattle have created binder proteins that strongly interact with 12 different protein targets.

Each binder was designed on a computer and then tested in bench experiments. The binders are all smaller than 65 amino acids in length, super stable (they hold their shape) and they bind their target with nanomolar or picomolar affinity. Most drugs used today bind in the nanomolar range; picomolar affinity indicates an even stronger interaction.

Designing binders on a computer, the researchers say, is much like taking on a difficult climbing wall that only has "a few good footholds or handholds distant from each other." Prior computational approaches to design binders focused "on routes involving these footholds/handholds, but this greatly limits the possibilities and there may be no way to connect them into a successful route."

To climb this hypothetical wall, then, the researchers first designed a huge amount of tiny proteins that each, individually, weakly interact with a small part of the target protein's surface. These little proteins are disconnected from one another, like points floating in space, and about one billion of them were designed for each target protein.

This step of the process is all about identification; pick out all the possible handholds and footholds up the climbing wall, no matter how poor they appear.

In the second step, protein backbones were designed to weave around each little protein, tying them together in three-dimensional space. The result is like a string of Christmas lights: each light weakly interacts with the target protein, and a cord holds them all together. The more lights that are roped together, the stronger they grab onto the target protein. In the paper, more than 84,000 protein scaffolds were tested, but only about 34,500 held their shape.

This step is all about testing; have thousands of climbers weave their way up the climbing wall, in unique ways, to find all the possible routes.

In the final step, the number of possible binders were narrowed down. If a similar design appeared on the computer many times, for example, it was assigned a higher score and prioritized it in later steps.

This step is all about validation; if multiple climbers make their way to the top of the wall using a similar route, then that route might be optimal.

These steps, together, offer a general solution for designing very strong binding proteins. Only a small fraction of the many thousands of generated binders actually bind to their protein target, though, so there is room for improvement. Still, the use of this technique for drug discovery — already the subject of a filed patent — seems imminent.

Read more at Nature.

(* = open access, † = review article)

*Ultrasound-controllable engineered bacteria for cancer immunotherapy. Abedi MH...Shapiro MG. Nature Communications. Link

An engineered prime editor with enhanced editing efficiency in plants. Zong Y...Gao C. Nature Biotechnology. Link

*Activating cryptic biosynthetic gene cluster through a CRISPR–Cas12a-mediated direct cloning approach. Liang M...Tan G. Nucleic Acids Research. Link

Other Papers

Basic Research

*Model-guided design of the diversity of a synthetic human gut community. Connors BM...Venturelli OS. bioRxiv (preprint). Link

*Periodic spatial patterning with a single morphogen. Wang S, Garcia-Ojalvo J & Elowitz MB. bioRxiv (preprint). Link

*Optogenetic control of the Bicoid morphogen reveals fast and slow modes of gap gene regulation. Singh AP...Toettcher JE. Cell Reports. Link


A Versatile Transcription Factor Biosensor System Responsive to Multiple Aromatic and Indole Inducers. Nasr MA...Kwan DH. ACS Synthetic Biology. Link


Design, mutate, screen: Multiplexed creation and arrayed screening of synchronized genetic clocks. Lezia A, Csicsery N & Hasty J. Cell Systems. Link

*Modular, robust and extendible multicellular circuit design in yeast. Carignano A...Klavins E. eLife. Link

*Cotranscriptionally encoded RNA strand displacement circuits. Schaffter SW & Strychalski EA. Science Advances. Link

†Designing Biological Circuits: From Principles to Applications. Chakraborty D, Rengaswamy R & Raman K. ACS Synthetic Biology. Link

*Identifying competition phenotypes in synthetic biochemical circuits. Al-Radhawi MA, Del Vecchio D & Sontag ED. bioRxiv (preprint). Link

Computation & Models

*Gene expression noise accelerates the evolution of a biological oscillator. Lin YT & Buchler NE. bioRxiv (preprint). Link

*Multicellular PI control for gene regulation in microbial consortia. Martinelli V...di Bernardo M. bioRxiv (preprint). Link

*The genetic and biochemical determinants of mRNA degradation rates in mammals. Agarwal V & Kelley D. bioRxiv (preprint). Link

*Using protein-per-mRNA differences among human tissues in codon optimization. Hernandez-Alias X...Schaefer MH. bioRxiv (preprint). Link

CRISPR & Genetic Engineering

*Guide RNAs containing universal bases enable Cas9/Cas12a recognition of polymorphic sequences. Krysler AR...Hubbard BP. Nature Communications. Link

*Harnessing DSB repair to promote efficient homology-dependent and -independent prime editing. Peterka M...Maresca M. Nature Communications. Link

*NT-CRISPR, combining natural transformation and CRISPR-Cas9 counterselection for markerless and scarless genome editing in Vibrio natriegens. Stukenberg D...Becker A. Communications Biology. Link

*Strategies to Identify and Edit Improvements in Synthetic Genome Segments Episomally. Rudolph A...Church G. bioRxiv (preprint). Link

*Targeting DNA polymerase to DNA double-strand breaks reduces DNA deletion size and increases templated insertions generated by CRISPR/Cas9. Yoo KW...Lu B. Nucleic Acids Research. Link

Medicine & Diagnostics

*Fast and sensitive detection of SARS-CoV-2 RNA using suboptimal protospacer adjacent motifs for Cas12a. Lu S...Yin H. Nature Biomedical Engineering. Link

*Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics. Leppek K...Das R. Nature Communications. Link

*Using CRISPR-Kill for organ specific cell elimination by cleavage of tandem repeats. Schindele A...Puchta H. Nature Communications. Link

*Sense-and-Respond Payload Delivery Using a Novel Antigen-Inducible Promoter Improves Suboptimal CAR-T Activation. Guo T, Ma D & Lu TK. ACS Synthetic Biology. Link

AAV-delivered suppressor tRNA overcomes a nonsense mutation in mice. Wang J...Wang D. Nature. Link

*Exon skipping induced by CRISPR-directed gene editing regulates the response to chemotherapy in non-small cell lung carcinoma cells. Banas K...Kmiec EB. Gene Therapy. Link

Orthogonally induced differentiation of stem cells for the programmatic patterning of vascularized organoids and bioprinted tissues. Skylar-Scott MA...Lewis JA. Nature Biomedical Engineering. Link

*Precise tumor immune rewiring via synthetic CRISPRa circuits gated by concurrent gain/loss of transcription factors. Wang Y...Liu J. Nature Communications. Link

Bioinstructive implantable scaffolds for rapid in vivo manufacture and release of CAR-T cells. Agarwalla P...Brudno Y. Nature Biotechnology. Link

*CRISPR activation screen identifies BCL-2 proteins and B3GNT2 as drivers of cancer resistance to T cell-mediated cytotoxicity. Joung J...Zhang F. Nature Communications. Link

Generation and characterization of hair-bearing skin organoids from human pluripotent stem cells. Lee J...Koehler KR. Nature Protocols. Link

Metabolic Engineering

*Design of stable and self-regulated microbial consortia for chemical synthesis. Li X...Yuan Q. Nature Communications. Link

Engineering the Lipid and Fatty Acid Metabolism in Yarrowia lipolytica for Sustainable Production of High Oleic Oils. Wang K...Ji X. ACS Synthetic Biology. Link

*Metabolic engineering strategies to produce medium-chain oleochemicals via acyl-ACP:CoA transacylase activity. Yan Q...Pfleger BF. Nature Communications. Link

*Biosynthesizing structurally diverse diols via a general route combining oxidative and reductive formations of OH-groups. Liu Y, Wang W & Zeng A. Nature Communications. Link


*A copper switch for inducing CRISPR/Cas9-based transcriptional activation tightly regulates gene expression in Nicotiana benthamiana. Garcia-Perez E...Vazquez-Vilar M. BMC Biotechnology. Link

*Optimization of CRISPR/LbCas12a-mediated gene editing in Arabidopsis. Zhang Q, Zhang Y & Chai Y. PLOS ONE. Link

*Plant-based expression and characterization of SARS-CoV-2 virus-like particles presenting a native spike protein. Jung J...Lomonossoff GP. Plant Biotechnology Journal. Link

Protein & Molecular Engineering

Charge Engineering Improves the Performance of Bst DNA Polymerase Fusions. Paik I, Bahdra S & Ellington AD. ACS Synthetic Biology. Link

*De novo design and directed folding of disulfide-bridged peptide heterodimers. Yao S...Wu C. Nature Communications. Link

Tools & Technology

*Optical control of protein delivery and partitioning in the nucleolus. Tan P...Zhou Y. Nucleic Acids Research. Link

A Temperature-Controlled Cell-Free Expression System by Dynamic Repressor. yang J, Wang C & Lu Y. ACS Synthetic Biology. Link

Fragment-Directed Random Mutagenesis by the Reverse Kunkel Method. Lin W...Mou KY. ACS Synthetic Biology. Link

*Expanding biochemical knowledge and illuminating metabolic dark matter with ATLASx. MohammadiPeyhani H...Hatzimanikatis V. Nature Communications. Link

*A toolkit for recombinant production of seven human EGF family growth factors in active conformation. Ferreira AS...Zanchin NIT. Scientific Reports. Link


*A Linear Programming-Based Strategy to Save Pipette Tips in Automated DNA Assembly. Sechkar K, Tuza ZA & Stan G. Synthetic Biology. Link

*†Towards synthetic diatoms: The Phaeodactylum tricornutum Pt-syn 1.0 project. Pampuch M, Walker EJL & Karas BJ. Current Opinion in Green and Sustainable Chemistry. Link

*The Space-Exposed Kombucha Microbial Community Member Komagataeibacter oboediens Showed Only Minor Changes in Its Genome After Reactivation on Earth. de Carvalho DS...Góes-Neto A. Frontiers in Microbiology. Link

Until next time,

— Niko // @NikoMcCarty // niko@codonmag.com