Computational Design of Potent and Selective d-Peptide Agonists of the Glucagon-like Peptide-2 Receptor.

Here, we designed three d-GLP-2 agonists that activated the glucagon-like peptide-2 receptor (GLP-2R) cyclic adenosine monophosphate (cAMP) accumulation without stimulating the glucagon-like peptide-1 receptor (GLP-1R). All the d-GLP-2 agonists increased the protein kinase B phosphorylated (p-AKT) expression levels in a time- and concentration-dependent manner in vitro. The most effective d-GLP-2 analogue boosted the AKT phosphorylation 2.

Disrupting the α-synuclein-ESCRT interaction with a peptide inhibitor mitigates neurodegeneration in preclinical models of Parkinson's disease.

Accumulation of α-synuclein into toxic oligomers or fibrils is implicated in dopaminergic neurodegeneration in Parkinson's disease. Here we performed a high-throughput, proteome-wide peptide screen to identify protein-protein interaction inhibitors that reduce α-synuclein oligomer levels and their associated cytotoxicity. We find that the most potent peptide inhibitor disrupts the direct interaction between the C-terminal region of α-synuclein and CHarged Multivesicular body Protein 2B (CHMP2B), a component of the Endosomal Sorting Complex Required for Transport-III (ESCRT-III).

A universal deep-learning model for zinc finger design enables transcription factor reprogramming.

CysHis zinc finger (ZF) domains engineered to bind specific target sequences in the genome provide an effective strategy for programmable regulation of gene expression, with many potential therapeutic applications. However, the structurally intricate engagement of ZF domains with DNA has made their design challenging. Here we describe the screening of 49 billion protein-DNA interactions and the development of a deep-learning model, ZFDesign, that solves ZF design for any genomic target.

Targeting the Receptor-Binding Motif of SARS-CoV-2 with D-Peptides Mimicking the ACE2 Binding Helix: Lessons for Inhibiting Omicron and Future Variants of Concern.

The COVID-19 pandemic continues to spread around the world, with several new variants emerging, particularly those of concern (VOCs). Omicron (B.1.

PepNN: a deep attention model for the identification of peptide binding sites.

Protein-peptide interactions play a fundamental role in many cellular processes, but remain underexplored experimentally and difficult to model computationally. Here, we present PepNN-Struct and PepNN-Seq, structure and sequence-based approaches for the prediction of peptide binding sites on a protein. A main difficulty for the prediction of peptide-protein interactions is the flexibility of peptides and their tendency to undergo conformational changes upon binding.

Computational Design of Potent D-Peptide Inhibitors of SARS-CoV-2.

Blocking the association between the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein receptor-binding domain (RBD) and the human angiotensin-converting enzyme 2 (ACE2) is an attractive therapeutic approach to prevent the virus from entering human cells. While antibodies and other modalities have been developed to this end, d-amino acid peptides offer unique advantages, including serum stability, low immunogenicity, and low cost of production. Here, we designed potent novel D-peptide inhibitors that mimic the ACE2 α1-binding helix by searching a mirror-image version of the PDB.

Rapid and accurate structure-based therapeutic peptide design using GPU accelerated thermodynamic integration.

Peptide-based therapeutics are an alternative to small molecule drugs as they offer superior specificity, lower toxicity, and easy synthesis. Here we present an approach that leverages the dramatic performance increase afforded by the recent arrival of GPU accelerated thermodynamic integration (TI). GPU TI facilitates very fast, highly accurate binding affinity optimization of peptides against therapeutic targets.

Method to generate highly stable D-amino acid analogs of bioactive helical peptides using a mirror image of the entire PDB.

Biologics are a rapidly growing class of therapeutics with many advantages over traditional small molecule drugs. A major obstacle to their development is that proteins and peptides are easily destroyed by proteases and, thus, typically have prohibitively short half-lives in human gut, plasma, and cells. One of the most effective ways to prevent degradation is to engineer analogs from dextrorotary (D)-amino acids, with up to 10-fold improvements in potency reported.

Generation and Validation of Intracellular Ubiquitin Variant Inhibitors for USP7 and USP10.

Post-translational modification of the p53 signaling pathway plays an important role in cell cycle progression and stress-induced apoptosis. Indeed, a large body of work has shown that dysregulation of p53 and its E3 ligase MDM2 by the ubiquitin-proteasome system (UPS) promotes carcinogenesis and malignant transformation. Thus, drug discovery efforts have focused on the restoration of wild-type p53 activity or inactivation of oncogenic mutant p53 by targeted inhibition of UPS components, particularly key deubiquitinases (DUBs) of the ubiquitin-specific protease (USP) class.

Strategies to Develop Inhibitors of Motif-Mediated Protein-Protein Interactions as Drug Leads.

Protein-protein interactions are fundamental for virtually all functions of the cell. A large fraction of these interactions involve short peptide motifs, and there has been increased interest in targeting them using peptide-based therapeutics. Peptides benefit from being specific, relatively safe, and easy to produce.

Protein engineering by highly parallel screening of computationally designed variants.

Current combinatorial selection strategies for protein engineering have been successful at generating binders against a range of targets; however, the combinatorial nature of the libraries and their vast undersampling of sequence space inherently limit these methods due to the difficulty in finely controlling protein properties of the engineered region. Meanwhile, great advances in computational protein design that can address these issues have largely been underutilized. We describe an integrated approach that computationally designs thousands of individual protein binders for high-throughput synthesis and selection to engineer high-affinity binders.

Pooled screening for antiproliferative inhibitors of protein-protein interactions.

Protein-protein interactions (PPIs) are emerging as a promising new class of drug targets. Here, we present a novel high-throughput approach to screen inhibitors of PPIs in cells. We designed a library of 50,000 human peptide-binding motifs and used a pooled lentiviral system to express them intracellularly and screen for their effects on cell proliferation.

Subcellular localization of coagulation factor II receptor-like 1 in neurons governs angiogenesis.

Neurons have an important role in retinal vascular development. Here we show that the G protein-coupled receptor (GPCR) coagulation factor II receptor-like 1 (F2rl1, previously known as Par2) is abundant in retinal ganglion cells and is associated with new blood vessel formation during retinal development and in ischemic retinopathy. After stimulation, F2rl1 in retinal ganglion cells translocates from the plasma membrane to the cell nucleus using a microtubule-dependent shuttle that requires sorting nexin 11 (Snx11).

Large-scale interaction profiling of PDZ domains through proteomic peptide-phage display using human and viral phage peptidomes.

The human proteome contains a plethora of short linear motifs (SLiMs) that serve as binding interfaces for modular protein domains. Such interactions are crucial for signaling and other cellular processes, but are difficult to detect because of their low to moderate affinities. Here we developed a dedicated approach, proteomic peptide-phage display (ProP-PD), to identify domain-SLiM interactions.

Caveolin-1 regulation of dynamin-dependent, raft-mediated endocytosis of cholera toxin-B sub-unit occurs independently of caveolae.

Ganglioside GM1-bound cholera toxin-B sub-unit (CT-b) enters the cell via clathrin-coated pits and dynamin-independent non-caveolar raft-dependent endocytosis. Caveolin-1 (Cav1), associated with caveolae formation, is a negative regulator of non-caveolar raft-dependent endocytosis. In mammary epithelial tumour cells deficient for Mgat5, Cav1 is stably expressed at levels below the threshold for caveolae formation, forming stable oligomerized Cav1 microdomains or scaffolds that were shown to suppress EGFR signalling and reduce the plasma membrane diffusion rate of both EGFR and CT-b.

Early effects of spinal cord transection on skeletal muscle properties.

Modulation of beta-adrenergic receptor (beta-AR) activity affects muscle mass and could have a role in the reduction of muscle mass observed following spinal cord transection (Tx). The aims of this study were to examine the early acute effects of Tx on muscle mass, total and myofibrillar protein concentrations, cytochrome c oxidase activity, and beta-AR density of skeletal muscle, to ascertain if any change in muscle properties could be related to beta-AR signalling events. Female Sprague-Dawley rats (n = 33; approximately 255 g) were randomly assigned to 4 experimental groups: control 4 d, control 8 d, Tx 4 d, and Tx 8 d.

Proangiogenic effects of protease-activated receptor 2 are tumor necrosis factor-alpha and consecutively Tie2 dependent.

Objective: Angiogenesis is essential physiologically in growth and pathologically in tumor development, chronic inflammatory disorders, and proliferative retinopathies. Activation of protease-activated receptor 2 (PAR2) leads to a proangiogenic response, but its mechanisms have yet to be specifically described. Here, we investigated the mode of action of PAR2 in retinal angiogenesis.

Apoptosis detected in the amygdala following myocardial infarction in the rat.

Background: Myocardial infarction (MI) contains a risk factor for developing episodes of Major Depressive Disorder (MDD). Apoptosis is commonly observed in the reperfused myocardial infarcted heart, and recent findings suggest the existence of apoptosis in MDD. Cytokines, which are released by ischemic myocardium and which may induce apoptosis, have been proposed as a possible cause for MDD.

Alterations of beta-adrenoceptor responsiveness in postischemic myocardium after 72 h of reperfusion.

To determine the effect of a completely developed reperfused myocardial infarction model on beta-adrenoceptor responsiveness, we induced a 90-min regional ischemia followed by 72 h of reperfusion in dog hearts. Regional myocardial blood flow was determined after 60 min of ischemia using radioactive microspheres. beta-adrenoceptor density was reduced in the ischemic endocardium (95+/-16 fmol/mg) and epicardium (160+/-13 fmol/mg) compared to the nonischemic region (304+/-21 fmol/mg).