The clinical outcome of SARS-CoV-2 infections, which can range from asymptomatic to lethal, is crucially shaped by the concentration of antiviral antibodies and by their affinity to their targets. However, the affinity of polyclonal antibody responses in plasma is difficult to measure. Here we used microfluidic antibody affinity profiling (MAAP) to determine the aggregate affinities and concentrations of anti–SARS-CoV-2 antibodies in plasma samples of 42 seropositive individuals, 19 of which were healthy donors, 20 displayed mild symptoms, and 3 were critically ill. We found that dissociation constants, Kd, of anti–receptor-binding domain antibodies spanned 2.5 orders of magnitude from sub-nanomolar to 43 nM. Using MAAP we found that antibodies of seropositive individuals induced the dissociation of pre-formed spike-ACE2 receptor complexes, which indicates that MAAP can be adapted as a complementary receptor competition assay. By comparison with cytopathic effect–based neutralisation assays, we show that MAAP can reliably predict the cellular neutralisation ability of sera, which may be an important consideration when selecting the most effective samples for therapeutic plasmapheresis and tracking the success of vaccinations.

T. improves ␤-cell mass and glucose disposal in streptozotocin-induced diabetic mice and activates cAMP/PKA/␤-catenin signaling in ␤-cells in vitro. Am J Physiol Endocrinol Metab 304: E1263-E1272, 2013. First published April 9, 2013 doi:10.1152/ajpendo.00600.2012.-Recent studies have demonstrated that the COOH-terminal fragment of the incretin hormone glucagonlike peptide-1 (GLP-1), a nonapeptide GLP-1(28 -36)amide, attenuates diabetes and hepatic steatosis in diet-induced obese mice. However, the effect of this nonapeptide in pancreatic ␤-cells remains largely unknown. Here, we show that in a streptozotocin-induced mouse diabetes model, GLP-1(28 -36)amide improved glucose disposal and increased pancreatic ␤-cell mass and ␤-cell proliferation. An in vitro investigation revealed that GLP-1(28 -36)amide stimulates ␤-catenin (␤-cat) Ser 675 phosphorylation in both the clonal INS-1 cell line and rat primary pancreatic islet cells. In INS-1 cells, the stimulation was accompanied by increased nuclear ␤-cat content. GLP-1(28 -36)amide was also shown to increase cellular cAMP levels, PKA enzymatic activity, and cAMP response element-binding protein (CREB) and cyclic AMP-dependent transcription factor-1 (ATF-1) phosphorylation. Furthermore, GLP-1(28 -36)amide treatment enhanced islet insulin secretion and increased the growth of INS-1 cells, which was associated with increased cyclin D1 expression. Finally, PKA inhibition attenuated the effect of GLP-1(28 -36)amide on ␤-cat Ser 675 phosphorylation and cyclin D1 expression in the INS-1 cell line. We have thus revealed the beneficial effect of GLP-1(28 -36)amide in pancreatic ␤-cells in vitro and in vivo. Our observations suggest that GLP-1(28 -36)amide may exert its effect through the PKA/␤-catenin signaling pathway.glucagon-like peptide-1; protein kinase A; Wnt; cAMP response element-binding protein; insulin; bromodeoxyuridine THE PROGLUCAGON GENE encodes both the pancreatic hormone glucagon and the gut incretin hormone glucagon-like peptide-1 (GLP-1) (15,17). The exploration of mechanisms underlying the function of GLP-1 and another incretin hormone, gastric inhibitory polypeptide (GIP), has led to the development of two categories of novel therapeutic agents, namely GLP-1 analogs and DPP-IV inhibitors, for diabetes and potentially its complications (11,12). In addition to targeting pancreatic ␤-cells, GLP-1 also exerts its function in many other organs or tissues (1,4,7,40). The GLP-1 receptor (GLP-1R)-deficient mouse line has been utilized as a powerful tool in studying the function of GLP-1 not only in pancreatic ␤-cells but also in a number of organs that are importantly involved in glucose disposal and metabolic homeostasis (7). However, studies with this mouse model, along with investigations with other tools, suggested that certain functions of GLP-1 may not be mediated by its canonical receptor GLP-1R, whereas a previously assumed inactive form of GLP-1, namely GLP-1(9 -36)amide, has the therapeutic potential in certain cardiovascular disorders (2, 25).Both G...

Parkinson’s disease is associated with the aberrant aggregation of α-synuclein. Although the causes of this process are still unclear, post-translational modifications of α-synuclein are likely to play a modulatory role. Since α-synuclein is constitutively N-terminally acetylated, we investigated how this post-translational modification alters the aggregation behavior of this protein. By applying a three-pronged aggregation kinetics approach, we observed that N-terminal acetylation results in a reduced rate of lipid-induced aggregation and slows down both elongation and fibril-catalyzed aggregate proliferation. An analysis of the amyloid fibrils produced by the aggregation process revealed different morphologies for the acetylated and non-acetylated forms in both lipid-induced aggregation and seed-induced aggregation assays. In addition, we found that fibrils formed by acetylated α-synuclein exhibit a lower β-sheet content. These findings indicate that N-terminal acetylation of α-synuclein alters its lipid-dependent aggregation behavior, reduces its rate of in vitro aggregation, and affects the structural properties of its fibrillar aggregates.

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A wide variety of oligomeric structures are formed during the aggregation of proteins associated with neurodegenerative diseases. Such soluble oligomers are believed to be key toxic species in the related disorders; therefore, identification of the structural determinants of toxicity is of upmost importance. Here, we analysed toxic oligomers of α-synuclein and its pathological variants in order to identify structural features that could be related to toxicity and found a novel structural polymorphism within G51D oligomers. These G51D oligomers can adopt a variety of β-sheet-rich structures with differing degrees of α-helical content, and the helical structural content of these oligomers correlates with the level of induced cellular dysfunction in SH-SY5Y cells. This structure–function relationship observed in α-synuclein oligomers thus presents the α-helical structure as another potential structural determinant that may be linked with cellular toxicity in amyloid-related proteins.