Biology subjects

Strasser, J.

Publications and source records attributed to Strasser, J..

6 recordsLinked to original sources

bioRxiv2026

Mycoplasma immunoglobulin binding protein universally binds human antibodies, thereby reducing Fab arm flexibility and displacing antigens in immune complexes

Mycoplasma immunoglobulin binding (MIB) protein plays a key role in immune evasion across several Mycoplasma species. MIB tightly interacts with the Fab domain of immunoglobulin G, inducing conformational changes that disrupt the antigen-binding site. Here, we reveal that MIB binds strongly and universally to all major isotypes and subclasses of human antibodies and can thus serve as a bait protein to efficiently deplete antibodies from serum. Our results demonstrate strong interactions between MIB and a diverse set of both recombinant monoclonal and endogenous polyclonal antibodies, the latter purified from serum or colostrum. All antibodies, including IgG and IgA monomers, J-chain coupled IgA dimers, and IgM pentamers, consistently bind two copies of MIB per antibody protomer. Using cross-linking mass spectrometry, we pinpoint that antibody-MIB interactions involve conserved regions of the variable and constant antibody domains, independent of isotype. Furthermore, we demonstrate that MIB not only disrupts the Fab-antigen interaction but also restricts Fab flexibility, as visualized by atomic force microscopy. Conversely, MIB does not affect Fc-mediated protein interactions, as exemplified by the successful reconstitution of a 36-subunit immune complex, (IgG)6-(MIB)12-C1q.

biochemistry↗

bioRxiv2025

A kinetic model of antigen-dependent IgG oligomerization and complement binding

The classical complement pathway (CCP), an important branch of the mammalian immune system, is initiated through multivalent binding of complement protein C1 to Immunoglobulin G (IgG) antibody oligomers assembled on the surface of pathogens, infected or malignant cells, culminating in the formation of the membrane attack complex (MAC) and subsequent cell lysis. IgG oligomers can further engage immune effector cells through Fc{gamma} receptors or complement receptors, facilitating antibody-dependent cellular cytotoxicity (ADCC) and phagocytosis (ADCP). Detailed knowledge of the factors that drive IgG oligomerization is thus vitally important to establish and improve IgG based therapies. We here focus on the kinetics of antigen-dependent IgG oligomerization and develop a comprehensive model capable of predicting oligomer formation as a function of IgG concentration, antigen density, IgG subclass and Fc point mutants, as well as the presence of Fc-binding and thus oligomerization-inhibiting factors such as staphylococcal protein A (SpA). We characterize the underlying molecular interactions in single molecule force spectroscopy (SMFS) and grating coupled interferometry (GCI) experiments. By fitting experimental data from high-speed atomic force microscopy (HS-AFM) experiments, we further quantify key rate constants and thermodynamic parameters, including free energy changes associated with oligomerization and apply the model to predict complement-mediated lysis in liposomal vesicle-based assays. The presented mechanistic framework may serve as a basis for optimizing antibody engineering and pharmacokinetic/pharmacodynamic modeling in the context of immunotherapies exploiting the CCP.

biophysics↗

bioRxiv2024

Complement activation by IgG subclasses is governed by their ability to oligomerize upon antigen binding

Complement activation through antibody-antigen complexes is crucial in various pathophysiological processes such as infections, inflammation, and autoimmunity, but is also utilized in immunotherapies to eliminate infectious agents, regulatory immune cells, or cancer cells. Although the tertiary structures of the four IgG antibody subclasses are largely identical, complement recruitment and further activation depend strongly on subclass, which is commonly explained by the respective affinity for C1, the first component of the classical complement pathway. Contradicting this established view, we here demonstrate that complement activation by different IgG subclasses is determined by their varying ability to form IgG oligomers on antigenic surfaces large enough to multivalently bind and activate C1. We directly visualize the resulting IgG oligomer structures and characterize their distribution by means of high-speed atomic force microscopy (HS-AFM), quantify their complement recruitment efficiency from quartz crystal microbalance (QCM) experiments, and characterize their ability to activate complement on tumor cell lines as well as in vesicle-based complement lysis assays. We present a mechanistic model of the multivalent interactions that govern C1 binding to IgG oligomers and use this model to extract affinities and kinetic rate constants from real-time interaction QCM data. Together, our detailed characterization yields a comprehensive view on the parameters that govern complement activation by the different IgG subclasses, which may inform the design of future antibody therapies.

immunology↗

bioRxiv2021

C1q binding to surface-bound IgG is stabilized by C1r2s2 proteases

Complement is an important effector mechanism for antibody-mediated clearance of infections and tumor cells. Upon binding to target cells, the antibodys constant (Fc) domain recruits complement component C1 to initiate a proteolytic cascade that generates lytic pores and stimulates phagocytosis. The C1 complex (C1qr2s2) consists of the large recognition protein C1q and a heterotetramer of proteases C1r and C1s (C1r2s2). While interactions between C1 and IgG-Fcs are believed to be mediated by the globular heads of C1q, we here find that C1r2s2 proteases affect the capacity of C1q to form an avid complex with surface-bound IgG molecules (on various DNP-coated surfaces and pathogenic Staphylococcus aureus). The extent to which C1r2s2 contribute to C1q-IgG stability strongly differs between human IgG subclasses. Using antibody engineering of monoclonal IgG we reveal that hexamer-enhancing mutations improve C1q-IgG stability, both in absence and presence of C1r2s2. In addition, hexamer-enhanced IgGs targeting S. aureus mediate improved complement-dependent phagocytosis by human neutrophils. Altogether, these molecular insights into complement binding to surface-bound IgGs could be important for optimal design of antibody therapies.

immunology↗

bioRxiv2020

Comprehensive label-free characterization of extracellular vesicles and their surface proteins

Interest in mesenchymal stem cell derived extracellular vesicles (MSC-EVs) as therapeutic agents has dramatically increased over the last decade. Preclinical studies show that MSC-EVs have anti-apoptotic and neuroprotective effects, boost wound healing, and improve the integration of allogeneic grafts through immunomodulation. Current approaches to the characterization and quality control of EV-based therapeutics include particle tracking techniques, Western blotting, and advanced cytometry, but standardized methods are lacking. In this study, we established and verified quartz crystal microbalance (QCM) as highly sensitive label-free immunosensing technique for characterizing clinically approved umbilical cord MSC-EVs enriched by tangential flow filtration and ultracentrifugation. Using QCM in conjunction with common characterization methods, we were able to specifically detect EVs via EV (CD9, CD63, CD81) and MSC (CD44, CD49e, CD73) markers and gauge their prevalence. Additionally, we characterized the topography and elasticity of these EVs by atomic force microscopy (AFM), enabling us to distinguish between EVs and non-vesicular particles (NVPs) in a therapeutic formulation. This measurement modality makes it possible to identify EV sub-fractions, discriminate between EVs and NVPs, and to characterize EV surface proteins, all with minimal sample preparation and using label-free measurement devices with low barriers of entry for labs looking to widen their spectrum of characterization techniques. Our combination of QCM with impedance measurement (QCM-I) and AFM measurements provides a robust multi-marker approach to the characterization of clinically approved EV formulations and opens the door to improved quality control.

molecular biology↗

bioRxiv2020

Staphylococcal protein A inhibits complement activation by interfering with IgG hexamer formation

IgG molecules are essential players in the human immune response against bacterial infections. An important effector of IgG-dependent immunity is the induction of complement activation, a reaction that triggers a variety of responses that help to kill bacteria. Antibody-dependent complement activation is promoted by the organization of target-bound IgGs into hexamers that are held together via noncovalent Fc-Fc interactions. Here we show that Staphylococcal protein A (SpA), an important virulence factor and vaccine candidate of Staphylococcus aureus, effectively blocks IgG hexamerization and subsequent complement activation. Using native mass spectrometry and high-speed atomic force microscopy, we demonstrate that SpA blocks IgG hexamerization through competitive binding to the Fc-Fc interaction interface on IgG monomers. In concordance, we show that SpA interferes with the formation of (IgG)6:C1q complexes and prevents downstream complement activation on the surface of S. aureus. Lastly, we demonstrate that IgG3 antibodies against S. aureus can potently induce complement activation even in the presence of SpA. Altogether, this study identifies SpA as an immune evasion protein that specifically blocks IgG hexamerization.

microbiology↗