Unconventional structure and mechanisms for membrane interaction and translocation of the NF-κB-targeting toxin AIP56

Bacterial AB toxins are secreted key virulence factors that are internalized by target cells through receptor-mediated endocytosis, translocating their enzymatic domain to the cytosol from endosomes (short-trip) or the endoplasmic reticulum (long-trip). To accomplish this, bacterial AB toxins evolved a multidomain structure organized into either a single polypeptide chain or non-covalently associated polypeptide chains. The prototypical short-trip single-chain toxin is characterized by a receptor-binding domain that confers cellular specificity and a translocation domain responsible for pore formation whereby the catalytic domain translocates to the cytosol in an endosomal acidification-dependent way. In this work, the determination of the three-dimensional structure of AIP56 shows that, instead of a two-domain organization suggested by previous studies, AIP56 has three-domains: a non-LEE encoded effector C (NleC)-like catalytic domain associated with a small middle domain that contains the linker-peptide, followed by the receptor-binding domain. In contrast to prototypical single-chain AB toxins, AIP56 does not comprise a typical structurally complex translocation domain; instead, the elements involved in translocation are scattered across its domains. Thus, the catalytic domain contains a helical hairpin that serves as a molecular switch for triggering the conformational changes necessary for membrane insertion only upon endosomal acidification, whereas the middle and receptor-binding domains are required for pore formation. © 2023, The Author(s).This work was supported by National funds through FCT under the project UIDB/04293/2020 and by FEDER funds through Programa Operacional Factores de Competitividade – COMPETE and by national funds through FCT – Fundação para a Ciência e a Tecnologia under the project PTDC/BIA-MIC/29910/2017 to N.M.S.S. A.d.V. was funded by Portuguese national funds through the FCT and, when eligible, by COMPETE 2020 FEDER funds, under the Scientific Employment Stimulus–Individual Call 2021.02251.CEECIND/CP1663/CT0016. We acknowledge access to the HTX crystallization facility (Proposal ID: BIOSTRUCTX_8167) and SOLEIL, ESRF and ALBA synchrotrons for provision of measurement time and thank their staff for help with data collection. The authors acknowledge the support of i3S Scientific Platforms (https://www.i3s.up.pt/scientific-platforms.php) Advanced Light Microscopy, member of the national infrastructure PPBI-Portuguese Platform of BioImaging (supported by POCI-01-0145-FEDER-022122), Animal Facility, Biochemical and Biophysical Technologies and X-ray Crystallography. A special thanks to Dr. Marc Graille and Dr. João Morais Cabral for constructive discussions in structural biology and Dr. Dimitri Panagiotis Papatheodorou for providing plasmid p327

Qualidade do melão cantaloupe cv. Hy-Mark minimamente processado e armazenado sob refrigeração Quality of cantaloupe melon cv. Hy-Mark minimally processed and refrigerated

A maioria das pesquisas em melão minimamente processado é concentrada no tipo 'cantaloupe', devido à sua importância no mercado internacional. O objetivo deste trabalho foi avaliar a qualidade do melão cantaloupe cv. Hy-Mark minimamente processado. Os frutos foram lavados e sanitizados, sendo em seguida cortados na forma de cubos, imersos em solução de CaCl2, acondicionados em embalagens flexíveis PET (tereftalato de polietileno) e armazenados a 4ºC por 9 dias. O delineamento foi conduzido inteiramente casualizado, com três repetições. A intervalos de três dias, amostras foram coletadas e analisadas quanto a coliformes totais e fecais, e contagem de bactérias aeróbias mesófilas e bolores e leveduras, pH, sólidos solúveis totais, acidez, vitamina C, açúcares redutores totais, atividade de água, firmeza e análise sensorial através do atributo de aceitação global. O melão minimamente processado apresentou boa estabilidade física, físico-química, microbiológica e sensorial, estimando-se em 9 dias a vida útil deste produto a 4ºC.<br>Most of the research into minimally processed melon is focused on cantaloupe melon, due to its importance in the international market. The objective of this work was to evaluate the quality of cantaloupe melon cv. Hy-Mark, minimally processed and refrigerated. Fruits were washed, sanitized, cut and imbedded in a calcium chloride solution and packed in PET and stored at 4ºC during 9 days. At three-day intervals samples were collected and analyzed for total and fecal coliforms, counting of aerobic mesophilic bacterias, molds and yeast, SS (soluble solids), TTA (total titrable acidity), firmness, pH, TRS (total reducing sugar), Aw, vitamin C, color and acceptability. The minimally processed melon showed good overall physical, pyisico-chemical, microbiological and sensorial stability during the storage period