Alpha-synuclein (αSyn) is a lipid-binding protein that assembles into amyloid fibrils in Parkinson's disease (PD). Lipid binding can accelerate the rate of αSyn fibril formation by stimulating primary nucleation. Recently, the lysosomal protein glucocerebrosidase (GCase) has been identified as a major risk factor in both genetic and sporadic PD. Using solution-state nuclear magnetic resonance spectroscopy, we observe that GCase binds to the C-terminal region of αSyn and attenuates the lipid-binding affinity of αSyn. Mechanistically, this interaction model involves competition between lipids and GCase for αSyn, with lipids and GCase preferentially interacting with the N- and C-terminal regions respectively, in a tug-of-war mechanism. Moreover, the addition of GCase to mature αSyn fibrils increases the NMR-visible population of flexible αSyn, indicating altered fibril-associated αSyn dynamics. Our findings provide a mechanistic explanation for the observed links between changes in GCase abundance and PD.
Current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are effective against severe disease and death, but do not prevent viral infections, probably due to the limited mucosal immunity induced by intramuscular administration of the vaccine. Fusion of SARS-CoV-2 subunit immunogens with a human IgG Fc backbone can be used as a mucosal vaccine but its effectiveness in delivery in animal models, and its immunogenicity and the vaccine-induced protection against viral infections requires further studies. Here we investigate a bivalent RBD-Fc vaccine that includes the spike receptor-binding domains (RBDs) of the ancestral and BQ.1.1 variant of SARS-CoV-2. Ex vivo fluorescent imaging demonstrates that this vaccine can be effectively delivered to the lungs of mice through intranasal administration, with enhancement of retention in the nasal cavity and lung parenchyma. In mice, the vaccine elicited potent and broad-spectrum antibody responses against different variants including KP.3 which could persist for at least 3 months after booster. Importantly, it was able to induce RBD-specific mucosal IgA responses. Further, heterologous intranasal immunisation with adeno-vectored Chadv1 and RBD-Fc elicited both potent neutralising antibody and T cell responses. Immunised BALB/c and K18-hACE2-transgenic mice were also protected against viral challenge of XBB.1 and viral transmission was effectively limited in hamsters through intranasal immunisation. This work thus demonstrates the potential of RBD-Fc antigens as mucosal vaccines for prevention of breakthrough infections and onward transmission. Moreover, Fc-fusion proteins can be used as an effective mucosal vaccine strategy which can be used either alone or in combination with other vaccine technology to constitute heterologous immunisations, enabling strong protection against SARS-CoV-2 and other respiratory viruses.
VCF files used and cited in the article: Identification of grapevine clones via high-throughput amplicon sequencing: a proof-of-concept study
Membrane-bound O-acyltransferases (MBOATs) are membrane-embedded enzymes that catalyze acyl chain transfer to a diverse group of substrates, including lipids, small molecules, and proteins. MBOATs share a conserved structural core, despite wide-ranging functional specificity across both prokaryotes and eukaryotes. The structural basis of catalytic specificity, regulation and interactions with the surrounding environment remain uncertain. Here, we combine comparative molecular dynamics (MD) simulations with bioinformatics to assess molecular and interactional divergence across the family. In simulations, MBOATs differentially distort the bilayer depending on their substrate type. Additionally, we identify lipid binding sites surrounding reactant gates in the surrounding membrane. Complementary bioinformatic analyses reveal a conserved role for re-entrant loop-2 in MBOAT fold stabilization and a key hydrogen bond bridging DGAT1 dimerization. Finally, we predict differences in MBOAT solvation and water gating properties. These data are pertinent to the design of MBOAT-specific inhibitors that encompass dynamic information within cellular mimetic environments.
