Molecular super-gluing: a straightforward tool for antibody labelling and its application to mycotoxin biosensing

Mycotoxins are low molecular weight toxic compounds, which can cause severe health problems in animals and humans. Immunoassays allow rapid, simple and cost-effective screening of mycotoxins. Sandwich assays with a direct readout provide great improvement in terms of selectivity and sensitivity, compared to the widely used competitive assay formats, for the analysis of low molecular weight molecules. In this work, we report a non-competitive fluorescence anti-immune complex (IC) immunoassay, based on the specific recognition of HT-2 toxin with a pair of recombinant antibody fragments, namely antigen-binding fragment (Fab) (anti-HT-2 (10) Fab) and single-chain variable fragment (scFv) (anti-IC HT-2 (10) scFv). The SpyTag and SpyCatcher glue proteins were applied for the first time as a bioconjugation tool for the analysis of mycotoxins. To this aim, a SpyTag-mScarlet-I (fluorescent protein) and scFv-SpyCatcher fusion proteins were constructed, produced and fused in situ during the assay by spontaneous Tag-Catcher binding. The assay showed an excellent sensitivity with an EC50 of 4.8 ± 0.4 ng mL−1 and a dynamic range from 1.7 ± 0.3 to 13 ± 2 ng mL−1, an inter-day reproducibility of 8.5% and a high selectivity towards HT-2 toxin without cross-reactivity with other Fusarium toxins. The bioassay was applied to the analysis of the toxin in an oat reference material and in oat samples, with a LOD of 0.6 µg kg−1, and the results were validated by analysing a certificate reference material and by HPLC–MS/MS. Graphical abstract

Implementing Zn2+ ion and pH-value control into artificial mussel glue proteins by abstracting a His-rich domain from preCollagen

A chemically activated mussel-inspired polymerization of a His-rich peptide, yielded artificial mussel glue proteins, where β-sheets can be triggered to mimic both adhesive motifs and cohesion control mechanisms of the mussel adhesive apparatus.

Structure and assembly model for the Trypanosoma cruzi 60S ribosomal subunit

Ribosomes of trypanosomatids, a family of protozoan parasites causing debilitating human diseases, possess multiply fragmented rRNAs that together are analogous to 28S rRNA, unusually large rRNA expansion segments, and r-protein variations compared with other eukaryotic ribosomes. To investigate the architecture of the trypanosomatid ribosomes, we determined the 2.5-Å structure of the Trypanosoma cruzi ribosome large subunit by single-particle cryo-EM. Examination of this structure and comparative analysis of the yeast ribosomal assembly pathway allowed us to develop a stepwise assembly model for the eight pieces of the large subunit rRNAs and a number of ancillary “glue” proteins. This model can be applied to the characterization of Trypanosoma brucei and Leishmania spp. ribosomes as well. Together with other details, our atomic-level structure may provide a foundation for structure-based design of antitrypanosome drugs.

Larval pupation site preference and its relationship to the glue proteins in a few species of Drosophila

The pupation site preference and the quantity of larval salivary gland secretion proteins (glue proteins) were analysed in 15 species of Drosophila belonging to 3 species groups. The results showed that even under constant environmental conditions, the larvae of different species prefer to pupate at different sites in various proportions. Three patterns of pupation site preference could be recognized on the basis of the preponderance of larvae pupating at different sites. The classification of different species of Drosophila under study into 3 pattern groups does not correspond with their taxonomic classification. The larvae of those species that synthesize large quantities of glue protein tend to pupate in the medium, while those synthesizing half the quantity of glue proteins pupate on the sides of the container (glass wall). The quantity of glue protein is not correlated with the size of the salivary glands. Key words : Drosophila, larval salivary glands, glue proteins, pupation site preference.

Genes from two intermoult puffs in Drosophila virilis polytene chromosomes are differentially transcribed during larval development

Genes from two Drosophila virilis intermoult puffs were isolated by microcloning. From puff 16A on the X-chromosome a 2.9 kb DNA fragment was obtained, which hybridizes with three transcripts. Two of them represent the mRNAs for larval glue proteins. They are found in different abundancies in third larval instar salivary glands, but also in minor amounts in midgut and in fat body. In puff 55E on chromosome III two genes were identified. They are transcribed exclusively in salivary glands during all three larval instars. Therefore, their products must be related to another gland-specific function, which is sustained throughout larval life.