PROTEO

Dairy white wastewater (WW), a by-product of industrial cleaning processes, contains residual milk proteins that can be enzymatically converted into bioactive peptides. In this study, WW proteins were hydrolyzed using four enzymes, pepsin, trypsin, thermolysin, and pronase E, for up to 240min, and the resulting hydrolysates were evaluated for ACE inhibition, DPP-IV inhibition, and antioxidant capacity. Thermolysin hydrolysates exhibited the strongest ACE inhibition, with IC50 values as low as 21.0μg protein/mL, whereas pepsin and pronase E hydrolysates showed DPP-IV inhibitory activities with IC50 values of 2.4-3.1mg protein/mL. Pepsin hydrolysates presented the highest antioxidant capacity, reaching 3.5μM Trolox equivalents/mg protein. LC-MS/MS analysis combined with multivariate statistics identified 60 discriminant peptides, including 17 peptides previously reported to possess antihypertensive, antidiabetic, and/or antioxidant bioactivities. Based on a combination of PLS-DA loadings, QSAR scores, novelty relative to known bioactive peptides and physicochemical diversity, 20 peptides were synthesized and validated experimentally. Several peptides such as LRF, QW, GAWY, PPF, GPIVL, and SFNPTQL exhibited potent inhibitory effects, with micromolar IC50 values for ACE and/or DPP-IV. In comparison to chemically synthesized ACE inhibitors like captopril, the most potent peptide, LRF, is five times more active on a molar basis (11.34μM vs 63.06μM). These findings demonstrate that WW is a promising source of multifunctional peptides and that integrating peptidomics with machine learning accelerates peptide discovery and validation.

IntroductionHigh-temperature stress imposes an energetic cost on plant growth and negatively impacts agricultural productivity. This stress rapidly triggers the activation of Heat shock Factor (HsF) proteins, a family of transcription factors that maintain proteostasis. The cryptochrome CRY1 can physically interact with HsFA1d proteins to facilitate nucleus translocation and the regulation of genes that contributes to stress tolerance.MethodsWe combined structural predictions with experimental testing using yeast-two-hybrid and bimolecular fluorescence complementation assays.ResultsWe confirm that CRY1 PHR domain interacts extensively with multiple HsF proteins through their HR-A region of the conserved oligomerization HR-A/B domain interface. This interaction partially relies on salt bridges provided by the N- and C-terminus of HR-A region and a conserved interface centered around W352 of CRY1. HsFA3 shows the strongest affinity to CRY1 in yeast-two-hybrid assays notably thanks to a glutamate residue that interacts with R211 and R435 of CRY1. Mutating equivalent residue positions within HsFA1e or HsFC1 to a glutamate increased their interaction to CRY1.DiscussionOverall, our analysis allowed the identification of mutant candidate that could be used in selection or bioengineering endeavors to improve thermal stress tolerance.

The islet amyloid polypeptide (IAPP) is a 37-residue peptide hormone secreted by pancreatic β-cells that is known to aggregate into amyloid fibrils. These fibrils accumulate in the pancreatic islets of individuals afflicted with type 2 diabetes and are implicated in β-cell dysfunction. Metal ions such as copper and zinc are known to modulate IAPP fibrillization, yet the role of metal-induced oxidative modifications in this process remains largely unexplored. This study examines the non-enzymatic post-translational oxidation of IAPP and its effects on aggregation using the biologically relevant Cu/O2/ascorbate system. Mass spectrometry identified residues within the amyloidogenic region (residues 20–29) as the primary targets of oxidation. These oxidative modifications impaired the formation of cross-β-sheet amyloid fibrils and promoted the accumulation of amorphous aggregates. The H18A IAPP derivative, lacking the key metal-binding histidine, was also examined to assess the impact of sequence variation on oxidation and aggregation. Copper-mediated oxidation of H18A resulted in a broader distribution of oxidation sites and impacts fibril formation. These findings provide preliminary mechanistic insights into copper-induced oxidation and its impact on IAPP aggregation pathways.

BackgroundINO80 and SWR1 are evolutionarily related ATP-dependent chromatin remodeling complexes that regulate the chromatin occupancy of the histone variant H2A.Z, playing critical roles in transcriptional regulation, genome replication, and DNA repair. While the H2A.Z-related functions of INO80 and SWR1 are well characterized in budding yeast and metazoans, much less is known about their composition and chromatin-targeting mechanisms outside of the Opisthokonts. We previously found that a distinct bromodomain-containing protein, IBD1, is involved in multiple chromatin-related complexes, including the SWR1-complex, in the ciliate protozoan Tetrahymena thermophila.ResultsHere, we report that a closely related bromodomain-containing protein, IBD2, functions as an acetyl lysine reader module within a putative INO80 complex. Through iterative proteomic analyses, we show that the Tetrahymena INO80 complex retains several conserved subunits found in its yeast and metazoan counterparts. In vitro binding assays reveal that recombinant IBD2 preferentially recognizes acetylated histone H3 tails. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) demonstrates that IBD2 is enriched near transcription start sites and promoter regions. Notably, the IBD1 and IBD2 genomic binding profiles strongly correlate with that of H2A.Z (Hv1), supporting their functional association with the SWRI- and INO80-complexes.ConclusionsTogether, our findings support a model in which H2A.Z chromatin dynamics are modulated by SWR1- and INO80-complexes that are differentially recruited to chromatin via distinct bromodomain proteins that recognize specific histone acetylation marks.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13072-025-00614-5.

Abstract Recent developments in DNA synthesis and sequencing have allowed the construction of comprehensive gene variant libraries and their functional analysis. Achieving high-replication and thorough mutation characterization remains technically and financially challenging for long genes. Here, we developed an efficient, affordable and scalable library construction approach that relies on low-cost DNA synthesis and standard cloning technologies, which will increase accessibility to systematic mutational studies and help advance the field of protein science.

Traditionally, eukaryotic mRNAs were perceived as inherently monocistronic. However, recent insights from ribosome profiling (Ribo-seq) and proteomics studies challenge this paradigm. These investigations reveal that, beyond the currently annotated reference proteins (RefProts), there exist additional proteins known as alternative proteins (AltProts) and small open reading frames derived microproteins encoded in regions of mRNAs previously considered untranslated or in non-coding transcripts. This experimental evidence broadens the spectrum of functional proteins within cells, tissues, and organs, potentially offering crucial insights into biological processes. Notably, a significant proportion of these newly identified AltProts and microproteins demonstrates localization in mitochondria, contributing to the functions of mitochondrial complexes. This review delves into the overlooked realm of the alternative proteome within mitochondria, exploring the role of nuclear or mitochondrial-genome-encoded AltProts and microproteins in physiological and pathological cellular processes.

Fluorohydrins - or β-fluorinated alcohols - and their fluorinated group derivatives are a biologically relevant class of compounds, with applications ranging from PET tracers to cancer therapeutics. Recent efforts have unlocked asymmetric access to these related motifs through reactions of carbonyls, alkenes, organoboranes, and epoxides or transformations such as cyclizations or ring expansions. The present work provides an overview of synthetic approaches to various fluorohydrins that have been explored in the past decade, as well as selected examples of these syntheses applied to medicinal chemistry.

Background/Objectives: During the pandemic, client-facing workers were perceived to be at greater risk of SARS-CoV-2 infection. This study investigated the risk factors for SARS-CoV-2 infection among a cohort of 304 retail workers in the Quebec City metropolitan area. Methods: After providing consent, participants were interviewed to gather information on demographic, socioeconomic, behavioural, and occupational variables. They were subsequently followed for up to five visits, scheduled every 12 ± 4 weeks. The study covered critical periods before and during the emergence of the Omicron variants and included retrospective reporting of COVID-19 symptoms and virus detection tests to capture the pandemic's early stages. Results: During the observation period, 173 (57%) participants experienced a first episode of COVID-19. Serological evidence of recent infection was detected in 160 participants (53%), while 117 (38%) reported a positive virus detection test. In adjusted analyses, risk factors for infection included younger age, a diagnosis of lung disease, longer weekly working hours, more frequent social gatherings, and having received fewer than three doses of vaccine. Notably, the increased risk associated with younger age and longer working hours was observed only after the relaxation of public health measures in the spring of 2022. Conclusions: These data suggest that during the early years of the pandemic when strict public health measures were in place, retail work was not a significant risk factor for SARS-CoV-2 infection in Quebec City metropolitan area. These findings highlight the complex dynamics of COVID-19 transmission and the effectiveness of workplace protective measures.

Abstract Alternative proteins (AltProts) represent a newly recognized class of biologically active proteins encoded from alternative open reading frames (AltORFs) within already annotated genes. This study focuses on theSLC35A4gene, which encodes both the reference protein SLC35A4 and the alternative protein AltSLC35A4. Using a combination of microscopy and biochemical analyses, we confirmed the presence of AltSLC35A4 in the inner mitochondrial membrane, resolving previous conflicting reports. Previous studies employing ribosome profiling have revealed that during oxidative stress induced by sodium arsenite, the reference coding sequence ofSLC35A4exhibits the largest increase in translational efficiency among all cellular mRNAs. Our results confirmed this translational upregulation, with the emergence of SLC35A4 protein isoforms during oxidative stress in an upstream ORF-dependent manner. Notably, the expression of AltSLC35A4 remained unchanged during oxidative stress. Knock- out of SLC35A4 enhanced sensitivity to oxidative stress in a rescuable manner, indicating a direct implication for SLC35A4 in stress resistance. In conclusion, our research provides compelling evidence for the functional significance of the dual-coding nature ofSLC35A4for resistance to oxidative stress and highlights the importance of considering AltProts in the functional study of eukaryotic genes.

Despite its broad application in in vitro studies, the application of targeted protein degradation (TPD) to animal models faces considerable challenges. Here, we develop inducible and cell-type specific TPD systems in mice using two degron systems: Oryza sativa TIR1F74G (OsTIR1)-auxin-inducible degron 2 (AID2) and human cereblon (hCRBN)-SALL4 degron (S4D). Efficient degradation of Satb1Venus protein by these systems recapitulates phenotypes observed in the Satb1-deficient mice. These TPD are successfully applied in both the fetal and neonatal stages. The OsTIR1-AID2 system proves to be effective for membrane proteins such as PD-1, emulating the effects of the anti-PD-1 antibody. Degradation of Bcl11b reveals a role of Bcl11b which was not characterized by the Cre-loxP system. Collectively, in vivo TPD technologies developed in this study enable inducible, temporal, and cell type-specific depletion of target proteins with high efficacy in mice. These technologies have a wide range of applications in the diverse fields of biological and medical research.