02 Oct 16:03
by Alugubelli, Y. R.
We have witnessed three coronavirus (CoV) outbreaks in the past two decades, including the COVID-19 pandemic caused by SARS-CoV-2. Main protease (MPro) is a highly conserved and essential protease that plays key roles in viral replication and pathogenesis among various CoVs, representing one of the most attractive drug targets for antiviral drug development. Traditional antiviral drug development strategies focus on the pursuit of high-affinity binding inhibitors against MPro. However, this approach often suffers from issues such as toxicity, drug resistance, and a lack of broad-spectrum efficacy. Targeted protein degradation represents a promising strategy for developing next-generation antiviral drugs to combat infectious diseases. Here we leverage the proteolysis targeting chimera (PROTAC) technology to develop a new class of small-molecule antivirals that induce the degradation of SARS-CoV-2 MPro. Our previously developed MPro inhibitors MPI8 and MPI29 were used as MPro ligands to conjugate a CRBN E3 ligand, leading to compounds that can both inhibit and degrade SARS-CoV-2 MPro. Among them, MDP2 was demonstrated to effectively reduce MPro protein levels in 293T cells (DC50 = 296 nM), relying on a time-dependent, CRBN-mediated, and proteasome-driven mechanism. Furthermore, MPD2 exhibited remarkable efficacy in diminishing MPro protein levels in SARS-CoV-2-infected A549-ACE2 cells, concurrently demonstrating potent anti-SARS-CoV-2 activity (EC50 = 492 nM). This proof-of-concept study highlights the potential of PROTAC-mediated targeted protein degradation of MPro as an innovative and promising approach for COVID-19 drug discovery.
02 Oct 16:03
by Schneider, H.
Proteolysis-targeting chimeras (PROTACs) have evolved in recent years from an academic idea to a therapeutic modality with more than 25 active clinical programs. However, achieving oral bioavailability and cell-type specificity remains a challenge, especially for PROTACs recruiting the von Hippel-Lindau (VHL) E3 ligase. Herein, we present an unprecedented, plug- and-play platform for VHL-recruiting PROTACs to overcome these limitations. Our platform allows for the generation of non-covalent antibody-PROTAC complexes within minutes and obviates the need for prior PROTAC modification, antibody-drug linker chemistry optimization or bioconjugation. Our technology relies on camelid-derived antibody domains (VHHs) which can easily be engineered into existing therapeutic antibody scaffolds. The resulting targeted, bispecific fusion proteins can be complexed with PROTACs at defined PROTAC-to-antibody ratios and have been termed PROxAb Shuttles. PROxAb Shuttles can prolong the half-life of PROTACs from hours to days, demonstrate anti-tumor efficacy in vivo and have the potential for reloading in vivo to further boost efficacy.
29 Sep 12:53
by Birk, A.
The mitochondrial membrane potential ({Delta}{Psi}m) is created by the accumulation of protons on an outer leaflet of the inner mitochondrial membrane and drives the synthesis of most cellular ATP, which is essential for cellular bioenergetics and survival. The {Delta}{Psi}m also facilitates the electrogenic transport of cations, such as Ca2+, and regulates generation of reactive oxygen species, which serves as a powerful bioenergetic and stress-signaling regulator. Proton trapping on the outer leaflet of the inner mitochondrial membrane of mitochondrial cristae could be controlled by cardiolipin when the local pH is above 8. However, there is presently no technology that effectively targets strong bases to cardiolipin.
We have developed a novel, high-density aromatic peptide (HDAP2) to preserve a proton gradient-driven potential in mitochondria by increasing proton trapping on cardiolipin (CL). HDAP2-induced formation of cardiolipin-HDAP2 complexes accumulated positive charges at the head of CL. The HDAP2-CL vesicles could accumulate the mitochondrial transmembrane potential probe, Tetramethylrhodamine (TMRM). This potential could be uncoupled with Carbonyl cyanide m-chlorophenylhydrazone (CCCP) and Dinitrophenol (DNP), indicating that an interaction of HDAP2 with CL could support a proton gradient-driven transmembrane potential.
We demonstrated that this novel, water-soluble peptide is cell-permeable, targets mitochondria without causing cell toxicity, and promotes cell survival during serum starvation. Importantly, the HDAP2-cardiolipin complex-mediated optimization of the proton gradient was supported by the ability of HDAP2 to prevent CCCP-mediated mitochondrial depolarization in ARPE-19 cells in a dose-dependent manner. Based on its mechanism of action, HDAP2 could promote cellular homeostasis, which would have broad clinical applicability for the prevention, recovery and reversal of many acute and chronic disease conditions, such as neurodegeneration, ischemia- reperfusion injury, and inflammation.
27 Sep 15:39
by Hsu, Y.-C.
Peptidoglycan (PGN), a net-like polymer constituted by muropeptides, provides protection for microorganisms and has been one of the major targets for antibiotics for decades. Researchers have explored host-microbiome interactions through PGN recognition systems and discovered key muropeptides modulating host responses. However, most common characterization techniques for muropeptides are labor-intensive and require manual analysis of mass spectra due to the complex cross-linked PGN structures. Each species has unique moiety modifications and inter-/intra-bridges, which further complicates the structural analysis of PGN. Here, we developed a high-throughput automated muropeptide analysis (HAMA) platform leveraging tandem mass spectrometry and in silico muropeptide MS/MS fragmentation matching to comprehensively identify muropeptide structures, quantify their abundance, and infer PGN cross-linking types. We demonstrated the effectiveness of the HAMA platform using well-characterized PGNs from E. coli and S. aureus and further applied it to common gut bacteria including species of Bifidobacterium, Bacteroides, Lactobacillus, Enterococcus, and Akkermansia. We thoroughly explored their PGN structures accurately identified muropeptide mono-/multi-mers, and even unambiguously discriminated the structural isomers via the HAMA platform. Furthermore, we found that the cell stiffness may be correlated to the compactness of the PGN structures through the length of interpeptide bridges or the site of transpeptidation within Bifidobacterium species. In summary, the HAMA framework exhibits an automated, intuitive, and accurate analysis of PGN compositions, which may serve as a potential tool to investigate the post-synthetic modifications of saccharides, the variation in interpeptide bridges, and the types of cross-linking within bacterial PGNs.
25 Sep 15:53
by Zheng LiuWenjie LiuWei WangYibao MaYufeng WangDavid L. DrumJinyang CaiHallie BlevinsEun LeeSyed ShahPaul B. FisherXinhui WangXianjun FangChunqing GuoXiang-Yang WangaDepartment of Human & Molecular Genetics, Virginia Commonwealth University School of Medicine, Richmond, VA 23298bDepartment of Biochemistry, Virginia Commonwealth University School of Medicine, Richmond, VA 23298cMassey Cancer Center, Virginia Commonwealth University School of Medicine, Richmond, VA 23298dDivision of Gastrointestinal and Oncologic Surgery, Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114eDepartment of Microbiology and Immunology, Virginia Commonwealth University School of Medicine, Richmond, VA 23298fHunter Holmes McGuire VA Medical Center, Richmond, VA 23249gInstitute of Molecular Medicine, Virginia Commonwealth University School of Medicine, Richmond, VA 23298
Proceedings of the National Academy of Sciences, Volume 120, Issue 39, September 2023.
25 Sep 15:52
by Morgane JaegerAmandine AnastasioLéa ChamySophie BrustleinRenaud VincentelliFabien DurbessonJulien GiganMorgane ThépautRémy CharMaud BoussandMathias LechelonRafael J. ArgüelloDidier MarguetHai-Tao HeRémi LasserreaAix Marseille Université, Centre National de la Recherche Scientifique, Institut National de la Santé et de la Recherche Médicale, Centre d’Immunologie de Marseille Luminy, Turing Center for Living Systems, 13 288 Marseille, FrancebAix Marseille Université, Institut National de la Santé et de la Recherche Médicale, Institut de neurobiologie de la Méditerranée, Turing Center for Living Systems, 13 273 Marseille, FrancecAix Marseille Université, Centre National de la Recherche Scientifique, Architecture et Fonction des Macromolécules Biologiques, 13 288 Marseille, France
Proceedings of the National Academy of Sciences, Volume 120, Issue 39, September 2023.
19 Sep 14:20
by Amélie Cabirol,
Silvia Moriano‐Gutierrez,
Philipp Engel
The gut microbiota of honeybees increases the abundance of several classes of neuroactive metabolites in the gut, the hemolymph, and the brain. Combining datasets from previous studies, we identify such metabolites and discuss how their deficiency might be responsible for the behavioral alterations reported in microbiota-free honeybees.
Abstract
Honey bees have emerged as a new model to study the gut–brain axis, as they exhibit complex social behaviors and cognitive abilities, while experiments with gnotobiotic bees have revealed that their gut microbiota alters both brain and behavioral phenotypes. Furthermore, while honey bee brain functions supporting a broad range of behaviors have been intensively studied for over 50 years, the gut microbiota of bees has been experimentally characterized only recently. Here, we combined six published datasets from metabolomic analyses to provide an overview of the neuroactive metabolites whose abundance in the gut, hemolymph and brain varies in presence of the gut microbiota. Such metabolites may either be produced by gut bacteria, released from the pollen grains during their decomposition by bacteria, or produced by other organs in response to different bacterial products. We describe the current state of knowledge regarding the impact of such metabolites on brain function and behavior and provide further hypotheses to explore in this emerging field of research.
19 Sep 14:19
by Mitch Leslie
Adding sugars to molecules that cause the body to attack itself shows promise in mice
12 Sep 20:52
by Agon Kokollari,
Marius Werner,
Christina Lindner,
Truc Lam Pham,
Franziska Thomas
Fast but not furious. A rapid on-resin N-formylation protocol for peptides was developed using formic acid, acetic anhydride, pyridine, and DMF. This method is simple in execution and provides near quantitative yield independent of peptide length and sequence.
Abstract
N-formylation is a common pre- and post-translational modification of the N-terminus or the lysine side chain of peptides and proteins that plays a role in the initiation of immune responses, gene expression, or epigenetics. Despite its high biological relevance, protocols for the chemical N-formylation of synthetic peptides are scarce. The few available methods are elaborate in their execution and the yields are highly sequence-dependent. We present a rapid, easy-to-use one-pot procedure that runs at room temperature and can be used to formylate protected peptides at both the N-terminus and the lysine side chain on the resin in near-quantitative yields. Only insensitive, storage-stable standard chemicals – formic acid, acetic anhydride, pyridine and DMF – are used. Formylation works for both short and long peptides of up to 34 amino acids and over the spectrum of canonical amino acids.