Structure-Based Druggability Assessment of the Mammalian Structural Proteome with Inclusion of Light Protein Flexibility

Loving, Kathryn; Lin, Andy; Cheng, Alan C.

doi:10.1371/journal.pcbi.1003741

Cited by 29 publications

(38 citation statements)

References 37 publications

(63 reference statements)

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“…2 C and S7 D ) and were previously studied using extensive molecular dynamics simulations in explicit solvent and Markov state models (6, 24). Moreover, both molecular dynamics simulations (23, 25, 48) and CryptoSite also successfully predicted known binding sites at difficult-to-drug protein-protein interaction interfaces, including in interleukin-2 (specificity of 79%), Bcl-X L (73%), FK506-binding protein (FKBP12; 73%), HPV regulatory protein E2 (50%), and cell division protein ZipA (60%) (Figs. 2 C , S7 E , and S8 C ).…”

Section: Resultsmentioning

confidence: 98%

“…In conclusion, CryptoSite is as accurate as approaches based on extensive molecular dynamics simulations, but significantly faster (a calculation on an average sized protein takes 1–2 days on our webserver) and completely automated. In comparison to approaches of similar efficiency (25, 28), CryptoSite is generally more accurate, particularly when the location of a cryptic site is buried in the apo state.…”

Section: Resultsmentioning

confidence: 99%

“…Currently, the only approaches to cryptic site discovery are exhaustive site-directed small-molecule tethering by experiment (20–22), long time-scale molecular dynamics simulations by computation (6, 8, 23, 24), flexible docking (25, 26), and computational tools for identification of small-molecule binding hot spots (10, 27–30). All of these approaches are time-consuming, expensive, and/or not always successful.…”

Section: Introductionmentioning

confidence: 99%

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CryptoSite: Expanding the Druggable Proteome by Characterization and Prediction of Cryptic Binding Sites

Cimermančič

Weinkam

Rettenmaier

et al. 2016

Journal of Molecular Biology

190

293

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Many proteins have small molecule-binding pockets that are not easily detectable in the ligand-free structures. These cryptic sites require a conformational change to become apparent; a cryptic site can therefore be defined as a site that forms a pocket in a holo structure, but not in the apo structure. Because many proteins appear to lack druggable pockets, understanding and accurately identifying cryptic sites could expand the set of drug targets. Previously, cryptic sites were identified experimentally by fragment-based ligand discovery, and computationally by long molecular dynamics simulations and fragment docking. Here, we begin by constructing a set of structurally defined apo-holo pairs with cryptic sites. Next, we comprehensively characterize the cryptic sites in terms of their sequence, structure, and dynamics attributes. We find that cryptic sites tend to be as conserved in evolution as traditional binding pockets, but are less hydrophobic and more flexible. Relying on this characterization, we use machine learning to predict cryptic sites with relatively high accuracy (for our benchmark, the true positive and false positive rates are 73% and 29%, respectively). We then predict cryptic sites in the entire structurally characterized human proteome (11,201 structures, covering 23% of all residues in the proteome). CryptoSite increases the size of the potentially “druggable” human proteome from ~40% to ~78% of disease-associated proteins. Finally, to demonstrate the utility of our approach in practice, we experimentally validate a cryptic site in protein tyrosine phosphatase 1B using a covalent ligand and NMR spectroscopy. The CryptoSite web server is available at http://salilab.org/cryptosite.

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Section: Resultsmentioning

confidence: 98%

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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CryptoSite: Expanding the Druggable Proteome by Characterization and Prediction of Cryptic Binding Sites

Cimermančič

Weinkam

Rettenmaier

et al. 2016

Journal of Molecular Biology

190

293

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“…The human and murine pockets have volumes of 170 Å 3 and 220 Å 3 , respectively. Notably, these pockets are comparable in size to other protein cavities with established small-molecule inhibitors (160-800 Å 3 ) (22)(23)(24)(25).…”

Section: Formation Of a Prominent Pocket In Human Pd-1 Upon Binding Pmentioning

confidence: 84%

“…A similar cavity forms in murine PD-1 upon binding of PD-L1 (20). Importantly, when murine PD-1 binds a different ligand, PD-L2 (21), this cavity extends ( Figures 1A-B) to a volume comparable to that occupied by established small-molecule inhibitors (22)(23)(24)(25). Unfortunately, this murine structure is insufficient to provide a structural model for the analogous pocket in the human PD-1/PD-L2 complex, as the human and murine PD-1 proteins share sequence identities of only 63% (26).…”

Section: Introductionmentioning

confidence: 86%

A high-affinity human PD-1/PD-L2 complex informs avenues for small-molecule immune checkpoint drug discovery

Tang

Kim

2019

Preprint

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Immune checkpoint blockade of programmed death-1 (PD-1) by monoclonal antibody drugs has delivered breakthroughs in the treatment of cancer. Nonetheless, smallmolecule PD-1 inhibitors could lead to increases in treatment efficacy, safety, and global access. While the ligand-binding surface of apo-PD-1 is relatively flat, it harbors a striking pocket in the murine PD-1/PD-L2 structure. An analogous pocket in human PD-1 may serve as a small-molecule drug target, but the structure of the human complex is unknown. Because the CC′ and FG loops in murine PD-1 adopt new conformations upon binding PD-L2, we hypothesized that mutations in these two loops could be coupled to pocket formation and alter PD-1's affinity for PD-L2. Here, we conducted deep mutational scanning in these loops and used yeast surface display to select for enhanced PD-L2 binding. A PD-1 variant with three substitutions binds PD-L2 with an affinity two orders of magnitude higher than that of the wild-type protein, permitting crystallization of the complex. We determined the X-ray crystal structures of the human triple-mutant PD-1/PD-L2 complex and the apo triple-mutant PD-1 variant at 2.0 Å and 1.2 Å resolution, respectively. Binding of PD-L2 is accompanied by formation of a prominent pocket in human PD-1, as well as substantial conformational changes in the CC′ and FG loops. The structure of the apo triple-mutant PD-1 shows that the CC′ loop adopts the ligand-bound conformation, providing support for allostery between the loop and pocket. This human PD-1/PD-L2 structure provide critical insights for the design and discovery of small-molecule PD-1 inhibitors. Significance StatementImmune checkpoint blockade of programmed death-1 (PD-1) by monoclonal antibody drugs has transformed the treatment of cancer. Small-molecule PD-1 drugs have the potential to offer increased efficacy, safety, and global access. Despite substantial efforts such small-molecule drugs have been out of reach. We identify a prominent pocket on the ligand-binding surface of human PD-1 that appears to be an attractive small-molecule drug target. The pocket forms when PD-1 is bound to one of its ligands, PD-L2. Our high-resolution crystal structure of the human PD-1/PD-L2 complex facilitates virtual drug-screening efforts and opens additional avenues for the design and discovery of small-molecule PD-1 inhibitors. Our work provides a strategy that may enable discovery of small-molecule inhibitors of other "undruggable" protein-protein interactions.binding ( Figure 2C). As a result, we obtained a PD-1 triple mutant (Figures S1F-G) that contains all three substitutions identified from the first library: N74G, T76P, and A132V. PD-1 loop variants showed increased binding affinity and association kinetics for PD-L2 and PD-L1To validate the detected enhancement in affinity, we recombinantly expressed and purified human PD-1 and the loop variants, as well as the human PD-L2 and PD-L1 ectodomain proteins. Using bio-layer interferometry, we compared the binding of PD-L2 to wild-type PD-1 and t...

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A Structural View on Druggability

Egner

Hillig

2020

Structural Biology in Drug Discovery

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Structure-Based Druggability Assessment of the Mammalian Structural Proteome with Inclusion of Light Protein Flexibility

Cited by 29 publications

References 37 publications

CryptoSite: Expanding the Druggable Proteome by Characterization and Prediction of Cryptic Binding Sites

CryptoSite: Expanding the Druggable Proteome by Characterization and Prediction of Cryptic Binding Sites

A high-affinity human PD-1/PD-L2 complex informs avenues for small-molecule immune checkpoint drug discovery

A Structural View on Druggability

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