Biology subjects

Pymm, P.

Publications and source records attributed to Pymm, P..

8 recordsLinked to original sources

bioRxiv2026

Nanobodies against Plasmodium vivax AMA1 block parasite invasion

Plasmodium vivax is the most widespread malaria species to infect human populations. P. vivax invades young red cells called reticulocytes to replicate and proliferate. An essential protein in parasite invasion is the Apical Membrane Antigen 1, PvAMA1, which forms the tight junction between parasite and red cell membrane through its interaction with Rhoptry Neck Protein 2, PvRON2. Here we report the identification of PvAMA1 nanobodies using phage display in combination with Sanger sequencing, next-generation sequencing and structure prediction tools. We show that these nanobodies bind with high affinities and across several different epitopes on PvAMA1. Several nanobodies recognize PvAMA1 in merozoites of late-stage schizonts and inhibit parasite invasion of red blood cells. These growth inhibitory nanobodies bind to conformational epitopes of PvAMA1 and most bind to a site distinct from a known potent human monoclonal antibody 826827. Crystal structures of two inhibitory nanobodies in complex with PvAMA1 show that they bind novel neutralizing epitopes which are outside the hydrophobic groove that contacts PvRON2. Furthermore, we show that two other PvAMA1 nanobodies also inhibit Plasmodium falciparum invasion. These results demonstrate the potential of PvAMA1 nanobodies to inhibit malaria parasite invasion across multiple Plasmodium species.

microbiology↗

bioRxiv2026

Cryo-EM structure of human LRRC15 reveals the basis of therapeutic antibody recognition

Leucine-rich repeat containing 15 (LRRC15) is a structural uncharacterised membrane protein, which is highly upregulated in cancer-associated fibroblasts and enriched in desmoplastic tumours including pancreatic, breast and head-and-neck cancers, where it is associated with therapy resistance. The LRRC15-targeting antibody-drug conjugate samrotamab vedotin has advanced into Phase I clinical evaluation, yet mechanistic understanding of how samrotamab engages its target has remained elusive in the absence of structural data. Here, using hydrogen-deuterium exchange mass spectrometry and cryo-electron microscopy, we define the samrotamab epitope on LRRC15 and determine the structure of the LRRC15-samrotamab complex to 2.6 [A] resolution - the first high-resolution structure of this receptor. Samrotamab engages a membrane-proximal epitope within the C-terminal leucine-rich repeat region, leaving the canonical concave surface fully exposed for potential interactions. The lateral binding mode of samrotamab and the open arc geometry of LRRC15 provide a structural rationale for why antibody occupancy may leave signalling-competent surfaces intact. Guided by these insights, we computationally designed minibinders targeting the concave surface, identifying multiple binders with nanomolar affinity. Together, our work establishes the first structural framework for LRRC15, defines the molecular basis of therapeutic antibody recognition, and identifies the concave surface as a promising target for next-generation LRRC15-directed therapeutics.

biochemistry↗

bioRxiv2026

Nanobodies against Plasmodium adhesins that block receptor engagement and malaria parasite invasion

Malaria is caused by Plasmodium parasites, and its clinical symptoms are a result of parasite invasion of red blood cells and the subsequent cycles of replication and proliferation. In human populations, Plasmodium vivax is responsible for the most widely distributed recurring malaria infections whereas Plasmodium falciparum inflicts the most mortality and morbidity. One well-characterized family of adhesins involved in red blood cell invasion is the reticulocyte-binding-like protein homolog family, known as the RBL superfamily which includes the PfRh family in P. falciparum and PvRBP family in P. vivax. Here we report a collection of nanobodies against three members of this adhesin family, PfRh5, PfRh4 and PvRBP2b. Nanobodies against these Plasmodium adhesins bind with high affinity across several epitopes and can block receptor engagement and inhibit parasite invasion of red blood cells. Using computational design, we generated stabilized PfRh4 variants that encompass the conserved scaffold present in the PfRh and PvRBP families of adhesins and show that several variants with improved expression retained binding to mouse monoclonal antibodies, nanobodies and Complement Receptor 1, the human receptor for PfRh4. We also observed that most of the inhibitory nanobodies against the three antigens recognized the conserved structural scaffold that define this family of adhesins. These results demonstrate the potential of nanobodies to block malaria parasite invasion into red blood cells.

microbiology↗

bioRxiv2025

Crystal structure and nanobodies against domain 3 of the malaria parasite fusogen Plasmodium falciparum HAP2

Malaria parasites are transmitted to humans through a bite from an infected female Anopheles mosquito. Within the mosquito midgut, malaria parasite gametes are activated and undergo fertilisation. If parasite fertilisation is perturbed, this stops the transmission of malaria parasites from mosquito to human. One proposed target of transmission-blocking interventions is Plasmodium falciparum fusogen PfHAP2, which is essential for gamete fusion during parasite fertilisation. However, to date, no monoclonal antibodies or structures of PfHAP2 have been generated. We have identified nanobodies that bind specifically to domain 3 of PfHAP2 with nanomolar affinities, two of which show some cross-species reactivity with HAP2 of other Plasmodium species. The crystal structure of one nanobody in complex with domain 3 of PfHAP2 provides the first structural insights into this transmission-blocking target in P. falciparum.

microbiology↗

bioRxiv2025

A novel chimeric coronavirus spike vaccine combining SARS-CoV-2 RBD and scaffold domains from HKU-1 elicits potent neutralising antibody responses

The receptor binding domain (RBD) of the SARS-CoV-2 spike is the major target for neutralising antibodies elicited by current vaccines. Using small domains such as the RBD as vaccine immunogens, however, may constrain the availability of CD4 T follicular helper (TFH) cells and impact immunogenicity. We engineered a novel chimeric trimeric RBD (CTR) glycoprotein, replacing the RBD of human coronavirus HKU-1 spike with SARS-CoV-2 RBD of either ancestral (WT) or Omicron BA.2 strains. This strategy maintains a native trimeric conformation of the RBD, while providing additional sources of CD4 T cell help via the HKU-1 spike scaffold. In C57BL/6 mice, CTR-BA.2 prime-boost vaccination elicited high anti-BA.2-RBD IgG and neutralising titres, matching responses in animals immunised with native SARS-CoV-2 spike proteins. GC B cells elicited by CTR-BA.2 were predominantly WT+/BA.2+ cross- reactive, and TFH cells predominantly recognised HKU-1 epitopes, demonstrating scaffold-directed T cell help. Macaques prime-boost immunised with CTR-WT similarly elicited high anti-RBD IgG, anti-spike IgG and neutralising responses, comparable to native spike-vaccinated animals. In draining lymph nodes of CTR-WT vaccinated macaques, RBD-specific GC B cells were present at elevated levels. In contrast to the murine studies, lymph node-draining TFH responses in macaques were broadly elicited against RBD, NTD/S2 or HKU-1-derived peptides. Although native SARS- CoV-2 spike was also highly immunogenic in animal models, our findings establish the chimeric glycoprotein design as a strategy to overcome the poor immunogenicity of the SARS-CoV-2 RBD by engaging CD4 TFH cells, while maintaining the ability to elicit protective neutralising responses. One sentence summaryA chimeric glycoprotein design preserves SARS-CoV-2 RBD antigenic conformation enabling elicitation of neutralising responses, while allowing recruitment of HKU-1 scaffold-directed CD4 helper responses to support the humoral response.

immunology↗

bioRxiv2024

Prediction of KIR3DL1/Human Leukocyte Antigen binding

KIR3DL1 is a polymorphic inhibitory Natural Killer (NK) cell receptor that recognizes Human Leukocyte Antigen (HLA) class I allotypes that contain the Bw4 motif. Structural analyses have shown that in addition to residues 77-83 that span the Bw4 motif, polymorphism at other sites throughout the HLA molecule can influence the interaction with KIR3DL1. Given the extensive polymorphism of both KIR3DL1 and HLA class I, we built a machine learning prediction model to describe the influence of allotypic variation on the binding of KIR3DL1 to HLA class I. Nine KIR3DL1 tetramers were screened for reactivity against a panel of HLA class I molecules which revealed different patterns of specificity for each KIR3DL1 allotype. Separate models were trained for each of KIR3DL1 allotypes based on the full amino sequence of exons 2 and 3 encoding the 1 and 2 domains of the class I HLA allotypes, the set of polymorphic positions that span the Bw4 motif, or the positions that encode 1 and 2 but exclude the connecting loops. The Multi-Label-Vector-Optimization (MLVO) model trained on all alpha helix positions performed best with AUC scores ranging from 0.74 to 0.974 for the 9 KIR3DL1 allotype models. We show that a binary division into binder and non-binder is not precise, and that intermediate levels exist. Using the same models, within the binder group, high- and low-binder categories can also be predicted, the regions in HLA affecting the high vs low binder being completely distinct from the classical Bw4 motif. We further show that these positions affect binding affinity in a nonadditive way and induce deviations from linear models used to predict interaction strength. We propose that this approach should be used in lieu of simpler binding models based on a single HLA motif.

immunology↗

bioRxiv2021

An iTSC-derived placental model of SARS-CoV-2 infection unveils ACE2-dependent susceptibility in syncytiotrophoblasts

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causing coronavirus disease 2019 (COVID-19) has caused a global health crisis. The primary site of infection is in the respiratory tract but the virus has been associated with a variety of complications involving the gastrointestinal and cardiovascular systems. Since the virus affects a variety of tissue types, there has been interest in understanding SARS-CoV-2 infection in early development and the placenta. ACE2 and TMPRSS2, two genes that are critical for SARS-CoV-2 virus entry are expressed in placenta-specific cell types including extravillous trophoblasts (EVTs) and especially, syncytiotrophoblasts (STs). The potential of SARS-CoV-2 to infect these placental cells and its effect on placental development and function is still unclear. Furthermore, it is crucial to understand the possible mechanism of vertical transmission of SARS-CoV-2 through the placenta. Here, we developed an in vitro model of SARS-CoV-2 infection of placental cell types using induced trophoblast stem cells (iTSCs). This model allowed us to show that STs but not EVTs are infected. Importantly, infected STs lack the expression of key differentiation genes, lack typically observed differentiated morphology and produce significantly lower human chorionic gonadotropin (HCG) compared to non-infected controls. We also show that an anti-ACE2 antibody prevents SARS-CoV-2 infection and restores normal ST differentiation and function. We highlight the establishment of a platform to study SARS-CoV-2 infection in early placental cell types, which will facilitate investigation of antiviral therapy to protect the placenta during early pregnancy and development.

cell biology↗

bioRxiv2020

Nanobody generation and structural characterization of Plasmodium falciparum 6-cysteine protein Pf12p

Surface-associated proteins play critical roles in the Plasmodium parasite life cycle and are major targets for vaccine development. The 6-cysteine (6-cys) protein family is expressed in a stage-specific manner throughout Plasmodium falciparum life cycle and characterized by the presence of 6-cys domains, which are {beta}-sandwich domains with conserved sets of disulfide bonds. Although several 6-cys family members have been implicated to play a role in sexual stages, mosquito transmission, evasion of the host immune response and host cell invasion, the precise function of many family members is still unknown and structural information is only available for four 6-cys proteins. Here, we present to the best of our knowledge, the first crystal structure of the 6-cys protein Pf12p determined at 2.8 [A] resolution. The monomeric molecule folds into two domains, D1 and D2, both of which adopt the canonical 6-cys domain fold. Although the structural fold is similar to that of Pf12, its paralog in P. falciparum, we show that Pf12p does not complex with Pf41, which is a known interaction partner of Pf12. We generated ten distinct Pf12p-specific nanobodies which map into two separate epitope groups; one group which binds within the D2 domain, while several members of the second group bind at the interface of the D1 and D2 domain of Pf12p. Characterization of the structural features of the 6-cys family and their associated nanobodies provide a framework for generating new tools to study the diverse functions of the 6-cys protein family in the Plasmodium life cycle.

microbiology↗