Biology subjects

Vanhoorelbeke, K.

Publications and source records attributed to Vanhoorelbeke, K..

4 recordsLinked to original sources

bioRxiv2025

Wrapping it Up: Structural Basis of ADAMTS13 Global Latency

ADAMTS13 is a critical enzyme responsible for cleaving ultra-large von Willebrand factor (VWF) multimers, thereby preventing the formation of microthrombi in the microvasculature. Dysfunction or deficiency of ADAMTS13 is associated with thrombotic thrombocytopenic purpura (TTP), a rare and life-threatening disorder characterized by microangiopathic hemolytic anemia and severe thrombocytopenia. Understanding the regulation of ADAMTS13 is essential for developing therapeutic interventions for TTP. ADAMTS13 exhibits two latency mechanisms: local latency within the metalloprotease (MP) domain and global latency that affects its overall conformation. Although the role of the two CUB domains in binding the Spacer module is known during global latency, the contributions of TSP7, TSP8, and the flexible Linker region (L3) between TSP8 and CUB1 have been repeatedly observed but never explained. Binding studies with monoclonal antibodies (mAbs) targeting the MP domain have revealed a cryptic epitope that becomes accessible only when ADAMTS13 is conformationally activated, and Spacer-CUB binding is disrupted. However, the mechanism behind this long-range effect has never been explained. We present a novel autoinhibition model of ADAMTS13, proposing that its distal domains directly occlude substrate binding sites, thereby modulating the enzymes activity. In this model, TSP7 and TSP8 bind directly to the MP module, while the Linker region between TSP8 and CUB1 acts as a pseudosubstrate, mimicking the natural VWF-A2 substrate and blocking the binding sites. Our results show that the CUB domains must depart from the tandem arrangement observed in the crystal structure and instead adopt refined binding positions. Further evidence comes from novel monoclonal antibodies targeting more cryptic epitopes on various ADAMTS13 domains that we have recently presented, as well as pH- and EDTA-dependent conformational changes observed by ELISA. A detailed molecular model is derived from extensive molecular simulations and mechanistically reconciles a wide range of previously disconnected experimental observations. This novel insight into the regulation of ADAMTS13 lays the groundwork for new therapeutic strategies against TTP. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=118 SRC="FIGDIR/small/689657v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@abed60org.highwire.dtl.DTLVardef@16d7470org.highwire.dtl.DTLVardef@f7a14aorg.highwire.dtl.DTLVardef@17e350f_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIAtomistic Structure of autoinhibited ADAMTS13-Del3To6 C_LIO_LIL3 linker (TSP8-CUB1) has a dual function: acting as a pseudosubstrate that blocks VWF-A2 binding sites and guiding CUB1-2 positioning C_LIO_LITSP7 and TSP8 bind at the MP domain to restrain activity C_LIO_LICUB1 and CUB2 domains are not a single functional block C_LI

molecular biology↗

bioRxiv2021

Anti-SARS-CoV-2 human antibodies retaining neutralizing activity against SARS-CoV-2 B.1.1.529 (omicron)

SARS-CoV-2 B.1.1.529, designated omicron, was recently identified as a new variant of concern by WHO and is rapidly replacing SARS-CoV-2 delta as the most dominant variant in many countries. Unfortunately, because of the high number of mutations present in the spike of SARS-CoV-2 omicron, most monoclonal antibodies (mAbs) currently approved for treatment of COVID-19 lose their in vitro neutralizing activity against this variant. We recently described a panel of human anti-SARS-CoV-2 mAbs that potently neutralize SARS-CoV-2 Wuhan, D614G and variants alpha, beta, gamma and delta. In this work, we evaluated our mAb panel for potential in vitro activity against SARS-CoV-2 delta and omicron. Three mAbs from our panel retain neutralizing activity against both delta and omicron, with mAb 3B8 still resulting in complete neutralization at a concentration as low as 0.02 g/ml for both variants. Overall, our data indicate that mAb 3B8 may have the potential to become a game-changer in the fight against the continuously evolving SARS-CoV-2.

immunology↗

bioRxiv2021

Potent neutralizing anti-SARS-CoV-2 human antibodies cure infection with SARS-CoV-2 variants in hamster model

Treatment with neutralizing monoclonal antibodies (mAbs) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributes to COVID-19 management. Unfortunately, SARS-CoV-2 variants can escape several of these recently approved mAbs, highlighting the need for additional discovery and development. In a convalescent COVID-19 patient, we identified six mAbs, classified in four epitope groups, that potently neutralized SARS-CoV-2 Wuhan, alpha, beta, gamma and delta infection in vitro. In hamsters, mAbs 3E6 and 3B8 potently cured infection with SARS-CoV-2 Wuhan, beta and delta when administered post-viral infection at 5 mg/kg. Even at 0.2 mg/kg, 3B8 still reduced viral titers. Intramuscular delivery of DNA-encoded 3B8 resulted in in vivo mAb production of median serum levels up to 90 g/ml, and protected hamsters against delta infection. Overall, our data mark 3B8 as a promising candidate against COVID-19, and highlight advances in both the identification and gene-based delivery of potent human mAbs.

immunology↗

bioRxiv2020

The GPIbα intracellular tail - role in transducing VWF- and Collagen/GPVI-mediated signaling

Synergy between GPIb and GPVI signaling machineries has been suggested previously, however its molecular mechanism remains unclear. We generated a novel GPIb transgenic mouse (GPIb{Delta}sig/{Delta}sig) by CRISPR-Cas9 technology to delete the last 24 residues of the GPIb intracellular tail important for VWF-mediated signaling. GPIb{Delta}sig/{Delta}sig platelets bound VWF normally under flow but formed fewer filopodia on VWF/botrocetin, demonstrating that the deleted region does not affect ligand binding but appreciably impairs VWF-dependent signaling. Notably, while haemostasis was normal in GPIb{Delta}sig/{Delta}sig mice, GPIb{Delta}sig/{Delta}sig platelets exhibited defective responses after collagen-related-peptide stimulation and formed smaller aggregates on collagen-coated microchannels at low and high shears. Flow assays performed with plasma-free blood or in the presence of IIb{beta}3-or GPVI-blockers suggested reduced IIb{beta}3 activation contributes to the phenotype of the GPIb{Delta}sig/{Delta}sig platelets. Together, these results reveal a new role for the intracellular tail of GPIb in transducing both VWF-GPIb and collagen-GPVI signaling events in platelets. Summary statementGPIb and GPVI are two key receptors on the platelet surface. Using a novel transgenic mouse (GPIb{Delta}sig/{Delta}sig) that lacks the last 24 amino acids of the GPIb intracellular tail, we demonstrate the importance of this region not only in transducing signals in response to GPIb binding to VWF, but also for collagen-GPVI-mediated platelet responses revealing previously underappreciated receptor crosstalk between GPIb and GPVI.

cell biology↗