Protein Science最新文献_第8页

Protein degradation of antizyme depends on the N-terminal degrons. 抗酶的蛋白质降解取决于 N 端脱胶子。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5199

Ju-Yi Hsieh, Pui-Ying Leong, Yi-Fang Yang, Yi-Liang Liu, Guang-Yaw Liu, Hui-Chih Hung

{"title":"Protein degradation of antizyme depends on the N-terminal degrons.","authors":"Ju-Yi Hsieh, Pui-Ying Leong, Yi-Fang Yang, Yi-Liang Liu, Guang-Yaw Liu, Hui-Chih Hung","doi":"10.1002/pro.5199","DOIUrl":"10.1002/pro.5199","url":null,"abstract":"Antizyme (AZ) is a regulatory protein that plays a crucial role in modulating the activity of ornithine decarboxylase (ODC), which is the initial and rate-limiting enzyme in the complex pathway of polyamine biosynthesis. AZ facilitates the swift degradation of ODC, thereby modulating the levels of cellular polyamines. This study unveils a new ubiquitin-independent mechanism for AZ degradation, emphasizing the essential role of N-terminal degrons. Contrary to traditional ubiquitin-dependent degradation, our findings reveal that AZ degradation is significantly influenced by its N-terminal region. By conducting a series of experiments, including in vitro degradation assays, cycloheximide chase experiments, differential scanning calorimetry, and measurement of cellular concentrations of polyamines, we demonstrate that N-terminal truncation significantly enhances AZ's stability and facilitates the reduction of polyamine levels by accelerating ODC degradation. The removal of the N-terminal portion of AZ results in a reduced degradation rate and enhanced thermal stability of the protein, leading to a more efficient inhibition of polyamine synthesis. These findings are corroborated by the analysis of AZ isoforms, AZ1, AZ2, and AZ3, which display differential degradation patterns based on the specific N-terminal segments. This substantiates a degradation mechanism driven by an intrinsically disordered N-terminal region acting as a degron, independent of lysine ubiquitination. These results underscore the significant regulatory function of the N-terminal domain in the activity of AZ and the maintenance of polyamine homeostasis.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5199"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11521938/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142547017","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Unique Fn3-like biosensor in σ^I/anti-σ^I factors for regulatory expression of major cellulosomal scaffoldins in Pseudobacteroides cellulosolvens. σI/抗σI因子中独特的Fn3类生物传感器，用于调控假杆菌（Pseudobacteroides cellulosolvens）中主要纤维素体支架蛋白的表达。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5193

Sheng Dong, Chao Chen, Jie Li, Ya-Jun Liu, Edward A Bayer, Raphael Lamed, Itzhak Mizrahi, Qiu Cui, Yingang Feng

{"title":"Unique Fn3-like biosensor in σI/anti-σI factors for regulatory expression of major cellulosomal scaffoldins in Pseudobacteroides cellulosolvens.","authors":"Sheng Dong, Chao Chen, Jie Li, Ya-Jun Liu, Edward A Bayer, Raphael Lamed, Itzhak Mizrahi, Qiu Cui, Yingang Feng","doi":"10.1002/pro.5193","DOIUrl":"10.1002/pro.5193","url":null,"abstract":"Lignocellulolytic clostridia employ multiple pairs of alternative σ/anti-σ (SigI/RsgI) factors to regulate cellulosomal components for substrate-specific degradation of cellulosic biomass. The current model has proposed that RsgIs use a sensor domain to bind specific extracellular lignocellulosic components and activate cognate SigIs to initiate expression of corresponding cellulosomal enzyme genes, while expression of scaffoldins can be initiated by several different SigIs. Pseudobacteroides cellulosolvens contains the most complex known cellulosome system and the highest number of SigI-RsgI regulons yet discovered. However, the function of many RsgI sensor domains and their relationship with the various enzyme types are not fully understood. Here, we report that RsgI4 from P. cellulosolvens employs a C-terminal module that bears distant similarity to the fibronectin type III (Fn3) domain and serves as the sensor domain. Substrate-binding analysis revealed that the Fn3-like domain of RsgI4 represents a novel carbohydrate-binding module (CBM) that binds to a wide range of polysaccharide types. Structure determination further revealed that the Fn3-like domain belongs to the type B group of CBMs with a predicted concave face for substrate binding. Promoter sequence analysis of cellulosomal genes revealed that SigI4 is responsible for cellulosomal regulation of major scaffoldins rather than enzymes, consistent with the broad substrate specificity of the RsgI4 sensor domain. Notably, scaffoldins are invariably required as cellulosome components regardless of the substrate type. These findings suggest that the intricate cellulosome system of P. cellulosolvens comprises a more elaborate regulation mechanism than other bacteria and thus expands the paradigm of cellulosome regulation.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5193"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11520246/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142522794","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Enhanced cellular death in liver and breast cancer cells by dual BET/BRPF1 inhibitors. BET/BRPF1 双重抑制剂可增强肝癌和乳腺癌细胞的细胞死亡。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5191

Giulia Cazzanelli, Andrea Dalle Vedove, Nicolò Sbardellati, Luca Valer, Amedeo Caflisch, Graziano Lolli

{"title":"Enhanced cellular death in liver and breast cancer cells by dual BET/BRPF1 inhibitors.","authors":"Giulia Cazzanelli, Andrea Dalle Vedove, Nicolò Sbardellati, Luca Valer, Amedeo Caflisch, Graziano Lolli","doi":"10.1002/pro.5191","DOIUrl":"10.1002/pro.5191","url":null,"abstract":"The acetylpyrrole scaffold is an acetylated lysine mimic that has been previously explored to develop bromodomain inhibitors. When tested on the hepatoma cell line Huh7 and the breast cancer cell line MDA-MB-231, a few compounds in our acetylpyrrole-thiazole library induced peculiar morphological changes, progressively causing cell death at increasing concentrations. Their evaluation on a panel of human bromodomains revealed concurrent inhibition of BRPF1 and BET bromodomains. To dissect the observed cellular effects, the acetylpyrrole derivatives were compared to JQ1 and GSK6853, chemical probes for the bromodomains of BET and BRPF1, respectively. The appearance of neurite-like extrusions, accompanied by βIII-tubulin overexpression, is caused by BET inhibition, with limited effect on cellular viability. Conversely, interference with BRPF1 induces cellular death but not phenotypic alterations. Combined treatment with JQ1 and GSK6853 showed additivity in reducing cellular viability, comparably to the acetylpyrrole-thiazole-based BET/BRPF1 inhibitors. In addition, we determined the crystallographic structures of the BRD4 and BRPF1 bromodomains in complex with the acetylpyrrole-thiazole compounds. The binding modes in the two bromodomains show similar interactions for the acetylpyrrole and different orientations of the moiety that point to the rim of the acetyl-lysine pocket.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5191"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11521936/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142547014","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Diversity and structural-functional insights of alpha-solenoid proteins. α-类烯醇蛋白的多样性和结构功能见解。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5189

Paula Nazarena Arrías, Zarifa Osmanli, Estefanía Peralta, Patricio Manuel Chinestrad, Alexander Miguel Monzon, Silvio C E Tosatto

{"title":"Diversity and structural-functional insights of alpha-solenoid proteins.","authors":"Paula Nazarena Arrías, Zarifa Osmanli, Estefanía Peralta, Patricio Manuel Chinestrad, Alexander Miguel Monzon, Silvio C E Tosatto","doi":"10.1002/pro.5189","DOIUrl":"10.1002/pro.5189","url":null,"abstract":"Alpha-solenoids are a significant and diverse subset of structured tandem repeat proteins (STRPs) that are important in various domains of life. This review examines their structural and functional diversity and highlights their role in critical cellular processes such as signaling, apoptosis, and transcriptional regulation. Alpha-solenoids can be classified into three geometric folds: low curvature, high curvature, and corkscrew, as well as eight subfolds: ankyrin repeats; Huntingtin, elongation factor 3, protein phosphatase 2A, and target of rapamycin; armadillo repeats; tetratricopeptide repeats; pentatricopeptide repeats; Pumilio repeats; transcription activator-like; and Sel-1 and Sel-1-like repeats. These subfolds represent distinct protein families with unique structural properties and functions, highlighting the versatility of alpha-solenoids. The review also discusses their association with disease, highlighting their potential as therapeutic targets and their role in protein design. Advances in state-of-the-art structure prediction methods provide new opportunities and challenges in the functional characterization and classification of this kind of fold, emphasizing the need for continued development of methods for their identification and proper data curation and deposition in the main databases.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5189"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11514114/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142506688","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Molecular and energetic analysis of the interaction and specificity of Maximin 3 with lipid membranes: In vitro and in silico assessments. Maximin 3 与脂质膜相互作用和特异性的分子和能量分析：体外和硅学评估。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5188

Pablo Luis Hernández-Adame, Brandt Bertrand, Martha Itzel Escamilla-Ruiz, Jaime Ruiz-García, Carlos Munoz-Garay

{"title":"Molecular and energetic analysis of the interaction and specificity of Maximin 3 with lipid membranes: In vitro and in silico assessments.","authors":"Pablo Luis Hernández-Adame, Brandt Bertrand, Martha Itzel Escamilla-Ruiz, Jaime Ruiz-García, Carlos Munoz-Garay","doi":"10.1002/pro.5188","DOIUrl":"10.1002/pro.5188","url":null,"abstract":"In this study, the interaction of antimicrobial peptide Maximin 3 (Max3) with three different lipid bilayer models was investigated to gain insight into its mechanism of action and membrane specificity. Bilayer perturbation assays using liposome calcein leakage dose-response curves revealed that Max3 is a selective membrane-active peptide. Dynamic light scattering recordings suggest that the peptide incorporates into the liposomal structure without producing a detergent effect. Langmuir monolayer compression assays confirmed the membrane inserting capacity of the peptide. Attenuated total reflection-Fourier transform infrared spectroscopy showed that the fingerprint signals of lipid phospholipid hydrophilic head groups and hydrophobic acyl chains are altered due to Max3-membrane interaction. On the other hand, all-atom molecular dynamics simulations (MDS) of the initial interaction with the membrane surface corroborated peptide-membrane selectivity. Peptide transmembrane MDS shed light on how the peptide differentially modifies lipid bilayer properties. Molecular mechanics Poisson-Boltzmann surface area calculations revealed a specific electrostatic interaction fingerprint of the peptide for each membrane model with which they were tested. The data generated from the in silico approach could account for some of the differences observed experimentally in the activity and selectivity of Max3.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5188"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11633330/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142547016","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Gating residues govern ligand unbinding kinetics from the buried cavity in HIF-2α PAS-B. 门控残基控制着配体从 HIF-2α PAS-B 的埋藏腔中解除结合的动力学。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5198

Marion L Silvestrini, Riccardo Solazzo, Soumendu Boral, Melanie J Cocco, Joseph D Closson, Matteo Masetti, Kevin H Gardner, Lillian T Chong

{"title":"Gating residues govern ligand unbinding kinetics from the buried cavity in HIF-2α PAS-B.","authors":"Marion L Silvestrini, Riccardo Solazzo, Soumendu Boral, Melanie J Cocco, Joseph D Closson, Matteo Masetti, Kevin H Gardner, Lillian T Chong","doi":"10.1002/pro.5198","DOIUrl":"10.1002/pro.5198","url":null,"abstract":"While transcription factors have been generally perceived as \"undruggable,\" an exception is the HIF-2 hypoxia-inducible transcription factor, which contains an internal cavity that is sufficiently large to accommodate a range of small-molecules, including the therapeutically used inhibitor belzutifan. Given the relatively long ligand residence times of these small molecules and the lack of any experimentally observed pathway connecting the cavity to solvent, there has been great interest in understanding how these drug ligands exit the buried receptor cavity. Here, we focus on the relevant PAS-B domain of hypoxia-inducible factor 2α (HIF-2α) and examine how one such small molecule (THS-017) exits from the buried cavity within this domain on the seconds-timescale using atomistic simulations and ZZ-exchange NMR. To enable the simulations, we applied the weighted ensemble path sampling strategy, which generates continuous pathways for a rare-event process [e.g., ligand (un)binding] with rigorous kinetics in orders of magnitude less computing time compared to conventional simulations. Results reveal the formation of an encounter complex intermediate and two distinct classes of pathways for ligand exit. Based on these pathways, we identified two pairs of conformational gating residues in the receptor: one for the major class (N288 and S304) and another for the minor class (L272 and M309). ZZ-exchange NMR validated the kinetic importance of N288 for ligand unbinding. Our results provide an ideal simulation dataset for rational manipulation of ligand unbinding kinetics.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5198"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11516114/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142522790","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

BEAN and HABAS: Polyphyletic insertions in the DNA-directed RNA polymerase. BEAN和HABAS：DNA定向RNA聚合酶中的多态插入。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5194

Claudia Alvarez-Carreño, Angela T Huynh, Anton S Petrov, Christine Orengo, Loren Dean Williams

{"title":"BEAN and HABAS: Polyphyletic insertions in the DNA-directed RNA polymerase.","authors":"Claudia Alvarez-Carreño, Angela T Huynh, Anton S Petrov, Christine Orengo, Loren Dean Williams","doi":"10.1002/pro.5194","DOIUrl":"10.1002/pro.5194","url":null,"abstract":"The β and β' subunits of the RNA polymerase (RNAP) are large proteins with complex multi-domain architectures that include several insertional domains. Here, we analyze the domain organizations of RNAP-β and RNAP-β' using sequence, experimentally determined structures and AlphaFold structure predictions. We observe that lineage-specific insertional domains in bacterial RNAP-β belong to a group that we call BEAN (broadly embedded annex). We observe that lineage-specific insertional domains in bacterial RNAP-β' belong to a group that we call HABAS (hammerhead/barrel-sandwich hybrid). The BEAN domain has a characteristic three-dimensional structure composed of two square bracket-like elements that are antiparallel relative to each other. The HABAS domain contains a four-stranded open β-sheet with a GD-box-like motif in one of the β-strands and the adjoining loop. The BEAN domain is inserted not only in the bacterial RNAP-β', but also in the archaeal version of universal ribosomal protein L10. The HABAS domain is inserted in several metabolic proteins. The phylogenetic distributions of bacterial lineage-specific insertional domains of β and β' subunits of RNAP follow the Tree of Life. The presence of insertional domains can help establish a relative timeline of events in the evolution of a protein because insertion is inferred to post-date the base domain. We discuss mechanisms that might account for the discovery of homologous insertional domains in non-equivalent locations in bacteria and archaea.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5194"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11515920/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142522789","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Tethered heme domains in a triheme cytochrome allow for increased electron transport distances. 三heme 细胞色素中的系链血红素域可增加电子传输距离。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5200

Benjamin W Nash, Tomás M Fernandes, Joshua A J Burton, Leonor Morgado, Jessica H van Wonderen, Dimitri A Svistunenko, Marcus J Edwards, Carlos A Salgueiro, Julea N Butt, Thomas A Clarke

{"title":"Tethered heme domains in a triheme cytochrome allow for increased electron transport distances.","authors":"Benjamin W Nash, Tomás M Fernandes, Joshua A J Burton, Leonor Morgado, Jessica H van Wonderen, Dimitri A Svistunenko, Marcus J Edwards, Carlos A Salgueiro, Julea N Butt, Thomas A Clarke","doi":"10.1002/pro.5200","DOIUrl":"10.1002/pro.5200","url":null,"abstract":"Decades of research describe myriad redox enzymes that contain cofactors arranged in tightly packed chains facilitating rapid and controlled intra-protein electron transfer. Many such enzymes participate in extracellular electron transfer (EET), a process which allows microorganisms to conserve energy in anoxic environments by exploiting mineral oxides and other extracellular substrates as terminal electron acceptors. In this work, we describe the properties of the triheme cytochrome PgcA from Geobacter sulfurreducens. PgcA has been shown to play an important role in EET but is unusual in containing three CXXCH heme binding motifs that are separated by repeated (PT)x motifs, suggested to enhance binding to mineral surfaces. Using a combination of structural, electrochemical, and biophysical techniques, we experimentally demonstrate that PgcA adopts numerous conformations stretching as far as 180 Å between the ends of domains I and III, without a tightly packed cofactor chain. Furthermore, we demonstrate a distinct role for its domain III as a mineral reductase that is recharged by domains I and II. These findings show PgcA to be the first of a new class of electron transfer proteins, with redox centers separated by some nanometers but tethered together by flexible linkers, facilitating electron transfer through a tethered diffusion mechanism rather than a fixed, closely packed electron transfer chain.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5200"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11520253/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142522793","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Expression and characterization of pantothenate energy-coupling factor transporters as an anti-infective drug target. 作为抗感染药物靶点的泛酸能量偶联因子转运体的表达和特征。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-11-01 DOI: 10.1002/pro.5195

Atanaz Shams, Spyridon Bousis, Eleonora Diamanti, Walid A M Elgaher, Lucie Zeimetz, Jörg Haupenthal, Dirk J Slotboom, Anna K H Hirsch

{"title":"Expression and characterization of pantothenate energy-coupling factor transporters as an anti-infective drug target.","authors":"Atanaz Shams, Spyridon Bousis, Eleonora Diamanti, Walid A M Elgaher, Lucie Zeimetz, Jörg Haupenthal, Dirk J Slotboom, Anna K H Hirsch","doi":"10.1002/pro.5195","DOIUrl":"10.1002/pro.5195","url":null,"abstract":"This study investigates the potential of energy-coupling factor (ECF) transporters as promising anti-infective targets to combat antimicrobial resistance (AMR). ECF transporters, a subclass of ATP-binding cassette (ABC) transporters, facilitate the uptake of B-vitamins across bacterial membranes by utilizing ATP as an energy source. Vitamins are essential cofactors for bacterial metabolism and growth, and they can either be synthesized de novo or absorbed from the environment. These transporters are considered promising drug targets, underscoring the need for further research to harness their medicinal potential and develop selective inhibitors that block vitamin uptake in bacteria. Herein, we focused on the ECF transporter for pantothenate (vitamin B5) from Streptococcus pneumoniae and the ECF transporter for folate (vitamin B9) from Lactobacillus delbrueckii as a reference protein. We also included the energizing module for pantothenate along with both full transporter complexes. Initially, we transformed and purified the transporters, followed by an assessment of their thermal stability under various buffer composition, pH, and salt concentrations. Additionally, we monitored the melting temperature over six days to confirm their stability for further assays. We then measured the binding affinities of six ECF inhibitors using surface plasmon resonance (SPR) and evaluated their inhibitory effects through in vitro assays, including bacterial growth assay, whole-cell uptake, and transport-activity assays. After determining cytotoxicity in two human cell lines, we established an in vivo infection model using Galleria mellonella larvae to further validate our findings.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 11","pages":"e5195"},"PeriodicalIF":4.5,"publicationDate":"2024-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11521937/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142547015","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Aggrescan4D: A comprehensive tool for pH-dependent analysis and engineering of protein aggregation propensity. Aggrescan4D：根据 pH 值分析和设计蛋白质聚集倾向的综合工具。

IF 4.5 3区生物学

Protein Science Pub Date : 2024-10-01 DOI: 10.1002/pro.5180

Mateusz Zalewski, Valentin Iglesias, Oriol Bárcenas, Salvador Ventura, Sebastian Kmiecik

{"title":"Aggrescan4D: A comprehensive tool for pH-dependent analysis and engineering of protein aggregation propensity.","authors":"Mateusz Zalewski, Valentin Iglesias, Oriol Bárcenas, Salvador Ventura, Sebastian Kmiecik","doi":"10.1002/pro.5180","DOIUrl":"10.1002/pro.5180","url":null,"abstract":"Aggrescan4D (A4D) is an advanced computational tool designed for predicting protein aggregation, leveraging structural information and the influence of pH. Building upon its predecessor, Aggrescan3D (A3D), A4D has undergone numerous enhancements aimed at assisting the improvement of protein solubility. This manuscript reviews A4D's updated functionalities and explains the fundamental principles behind its pH-dependent calculations. Additionally, it presents an antibody case study to evaluate its performance in comparison with other structure-based predictors. Notably, A4D integrates advanced protein engineering protocols with pH-dependent calculations, enhancing its utility in advising solubility-enhancing mutations. A4D considers the impact of structural flexibility on aggregation propensities, and includes a large set of precalculated predictions. These capabilities should help to open new avenues for both understanding and managing protein aggregation. A4D is accessible through a dedicated web server at https://biocomp.chem.uw.edu.pl/a4d/.","PeriodicalId":20761,"journal":{"name":"Protein Science","volume":"33 10","pages":"e5180"},"PeriodicalIF":4.5,"publicationDate":"2024-10-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11425640/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142352674","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0