Protein-protein interactions (PPIs) are well-established as a class of promising drug targets for their implications in a wide range of biological processes. However, drug development toward PPIs is inevitably hampered by their flat and wide interfaces, which generally lack suitable pockets for ligand binding, rendering most PPI systems "undruggable." Here, we summarized drug design strategies for developing peptide-based PPI inhibitors. Importantly, several quintessential examples toward well-established PPI targets such as Bcl-2 family members, p53-MDM2, as well as APC-Asef are presented to illustrate the detailed schemes for peptide-based PPI inhibitor development and optimizations. This review supplies a comprehensive overview of recent progresses in drug discovery targeting PPIs through peptides or peptidomimetics, and will shed light on future therapeutic agent development toward the historically "intractable" PPI systems.

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Rational Design of Peptide-Based Inhibitors Disrupting Protein-Protein Interactions

Class B G protein-coupled receptors (GPCRs) are important targets in the treatment of metabolic syndrome and diabetes. Although multiple structures of class B GPCRs-G protein complexes have been elucidated, the detailed activation mechanism of the receptors remains unclear. Here, we combine Gaussian accelerated molecular dynamics simulations and Markov state models (MSM) to investigate the activation mechanism of a canonical class B GPCR, human glucagon receptor-GCGR, including the negative allosteric modulator-bound inactive state, the agonist glucagon-bound active state, and both glucagon- and Gs-bound fully active state. The free-energy landscapes of GCGR show the conformational ensemble consisting of three activation-associated states: inactive, active, and fully active. The structural analysis indicates the high dynamics of GCGR upon glucagon binding with both active and inactive conformations in the ensemble. Significantly, the H8 and TM6 exhibits distinct features from the inactive to the active states. The additional simulations demonstrate the role of H8 in the recruitment of Gs. Gs binding presents a crucial function of stabilizing the glucagon binding site and MSM highlights the absolute requirement of Gs to help the GCGR reach the fully active state. Together, our results reveal the detailed activation mechanism of GCGR from the view of conformational dynamics. 

https://doi.org/10.1016/j.csbj.2022.01.015

Delineating the activation mechanism and conformational landscape of a class B G protein-coupled receptor glucagon receptor

Dual-targeting therapeutics by coadministration of allosteric and orthosteric drugs is drawing increased attention as a revolutionary strategy for overcoming the drug-resistance problems. It was further observed that the occupation of orthosteric sites by therapeutics agents has the potential to enhance allosteric ligand binding, which leads to improved potency of allosteric drugs. Epidermal growth factor receptor (EGFR), as one of the most critical anti-cancer targets belonging to the receptor tyrosine kinase family, represents a quintessential example. It was revealed that osimertinib, an ATP-competitive covalent EGFR inhibitor, remarkably enhanced the affinity of a recently developed allosteric inhibitor JBJ-04-125-02 for EGFRL858R/T790M. Here, we utilized extensive large-scale molecular dynamics simulations and the reversed allosteric communication to untangle the detailed molecular underpinning, in which occupation of osimertinib at the orthosteric site altered the overall conformational ensemble of EGFR mutant and reshaped the allosteric site via long-distance signaling. A unique intermediate state resembling the active conformation was identified, which was further stabilized by osimertinib loading. Based on the allosteric communication pathway, we predicted a novel allosteric site positioned around K867, E868, H893, and K960 within the intermediate state. Its correlation with the orthosteric site was validated by both structural and energetic analysis, and its low sequence conservation indicated the potential for selective targeting across the human kinome. Together, these findings not only provided a mechanistic basis for future clinical application of the dual-targeting therapeutics, but also explored an innovative perception of allosteric inhibition of tyrosine kinase signaling.

Untangling Dual-Targeting Therapeutic Mechanism of Epidermal Growth Factor Receptor (EGFR) Based on Reversed Allosteric Communication.

Atomic-scale insights into allosteric inhibition and evolutional rescue mechanism of Streptococcus thermophilus Cas9 by the anti-CRISPR protein AcrIIA6

Conformational transformation of switch domains in GDP/K-Ras induced by G13 mutants: An investigation through Gaussian accelerated molecular dynamics simulations and principal component analysis.

Ras undergoes interconversion between the active GTP-bound state and the inactive GDP-bound state This GTPase cycle, which controls the activities of Ras, is accelerated by Ras GTPase-activating proteins (GAPs) and guanine nucleotide exchange factors (SOS) Oncogenic Ras mutations could affect the GTPase cycle and impair Ras functions Additionally, Src-induced K-Ras Y32/64 dual phosphorylation has been reported to disrupt GTPase cycle and hinder Ras downstream signaling However, the underlying mechanisms remain unclear To address this, we performed molecular dynamics simulations (~30 μs in total) on unphosphorylated and phosphorylated K-Ras4B in GTP- and GDP-bound states, and on their complexes with GTPase cycle regulators (GAP and SOS) and the effector protein Raf We found that K-Ras4B dual phosphorylation mainly alters the conformation at the nucleotide binding site and creates disorder at the catalytic site, resulting in the enlargement of GDP binding pocket and the retard of Ras-GTP intrinsic hydrolysis We observed phosphorylation-induced shift in the distribution of Ras-GTP inactive-active sub-states and recognized potential druggable pockets in the phosphorylated Ras-GTP Moreover, decreased catalytic competence or signal delivery abilities due to reduced binding affinities and/or distorted catalytic conformations of GAP, SOS and Raf were observed In addition, the allosteric pathway from Ras/Raf interface to the distal Raf L4 loop was compromised by Ras phosphorylation These results reveal the mechanisms by which phosphorylation influences the intrinsic or GAP/SOS catalyzed transformations between GTP- and GDP-bound states of Ras and its signal transduction to Raf Our findings project Ras phosphorylation as a target for cancer drug discovery

Dong Ji

Papers

Mechanistic insights into the effect of phosphorylation on Ras conformational dynamics and its interactions with cell signaling proteins