BiochimiePub Date : 2023-12-28DOI: 10.1016/j.biochi.2023.12.010
Takayuki K. Nemoto , Haruka Nishimata , Kana Shirakura , Yuko Ohara-Nemoto
{"title":"Potential elevation of exopeptidase activity of Glu-specific endopeptidase I/GluV8 mediated by hydrophobic P1′-position amino acid residue","authors":"Takayuki K. Nemoto , Haruka Nishimata , Kana Shirakura , Yuko Ohara-Nemoto","doi":"10.1016/j.biochi.2023.12.010","DOIUrl":"10.1016/j.biochi.2023.12.010","url":null,"abstract":"<div><p><span><span>We recently reported that the activities of dipeptidyl-peptidase (DPP)7 and DPP11, S46-family exopeptidases were significantly elevated by the presence of prime-side </span>amino acid residues of substrates caused by an increase in </span><em>k</em><sub>cat</sub><span> [Ohara-Nemoto Y. et al., J Biol Chem 298(3):101585. doi: 10.1016/j.jbc.2022]. In the present study, the effects of prime-side residues on Glu-specific endopeptidase I/GluV8 from </span><span><em>Staphylococcus aureus</em></span><span> were investigated using a two-step cleavage method with tetrapeptidyl-methycoumaryl-7-amide (MCA) carrying P2- to P2′-position residues coupled with DPP11 as the second enzyme<span>. GluV8 showed maximal activity toward benzyloxycarbonyl (Z)-LLE-MCA, while the effects of hydrolysis of substrates one residue shorter</span></span><u>,</u><span> such as acetyl (Ac)-Val-Glu- and Leu-Glu-MCA, were negligible. Nevertheless, activity towards Ac-VE-|-ID-MCA, a substrate carrying P1′ and P2′ residues, emerged and reached a level 44 % of that for Z-LLE-MCA. Among 11 Ac-HAXD-MCA (X is a varied amino acid), the highest level of activity enhancement was achieved with P1′-Leu and Ile, followed by Phe, Val, Ser, Tyr, and Ala, while Gly and Lys showed scant effects. This activation order was in parallel with the hydrophobicity indexes of these amino acids. The prime-side residues increased </span><em>k</em><sub>cat</sub>/<em>K</em><sub>M</sub> primarily through a maximum 500-fold elevation of <em>k</em><sub>cat</sub><span> as well as S46-family exopeptidases. The MEROPS substrate database also indicates a close relationship between activity and hydrophobicity of the P1′ residues in 93 N-terminal-truncated substrates, though no correlation was observed among all 4328 GluV8 entities examined. Taken together, these results are the first to demonstrate N-terminal exopeptidase activity of GluV8, considered to be prompted by hydrophobic P1′ amino acid residues.</span></p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139068881","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-27DOI: 10.1016/j.biochi.2023.12.008
Francisco Alejandro Lagunas-Rangel
{"title":"Naked mole-rat hyaluronan","authors":"Francisco Alejandro Lagunas-Rangel","doi":"10.1016/j.biochi.2023.12.008","DOIUrl":"10.1016/j.biochi.2023.12.008","url":null,"abstract":"<div><p><span><span><span><span>Naked mole rats (NMRs) are renowned for their exceptional longevity and remarkable maintenance of health throughout their lifetime. Their subterranean lifestyle has led to adaptations that have resulted in elevated levels of a very high molecular weight </span>hyaluronan in their tissues. Hyaluronan, a </span>glycosaminoglycan, is a key component of the extracellular matrix, which plays a critical role in maintaining tissue structure and regulating </span>cell signaling pathways. This phenomenon in NMRs is attributed to a higher processing and production capacity by some of their </span>hyaluronan synthases<span><span>, along with lower degradation by certain hyaluronidases. Furthermore, this adaptation indirectly confers several advantages to NMRs, such as the preservation of skin elasticity and youthful appearance, accelerated wound healing, protection against </span>oxidative stress<span>, and resistance to conditions such as cancer and arthritis, largely attributable to CD44 signaling and other intricate mechanisms. Thus, the main objective of this study was to conduct a comprehensive study of the distinctive features of NMR hyaluronan, particularly emphasizing the currently known molecular mechanisms that contribute to its beneficial properties. Furthermore, this research delves into the potential applications of NMR hyaluronan in both cosmetic and therapeutic fields, as well as the challenges involved.</span></span></p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139055543","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-27DOI: 10.1016/j.biochi.2023.12.007
Jorgaq Pata , Alexis Moreno , Benjamin Wiseman , Sandrine Magnard , Idriss Lehlali , Marie Dujardin , Atanu Banerjee , Martin Högbom , Ahcène Boumendjel , Vincent Chaptal , Rajendra Prasad , Pierre Falson
{"title":"Purification and characterization of Cdr1, the drug-efflux pump conferring azole resistance in Candida species","authors":"Jorgaq Pata , Alexis Moreno , Benjamin Wiseman , Sandrine Magnard , Idriss Lehlali , Marie Dujardin , Atanu Banerjee , Martin Högbom , Ahcène Boumendjel , Vincent Chaptal , Rajendra Prasad , Pierre Falson","doi":"10.1016/j.biochi.2023.12.007","DOIUrl":"10.1016/j.biochi.2023.12.007","url":null,"abstract":"<div><p><span><em>Candida albicans</em></span> and <em>C. glabrata</em><span> express exporters of the ATP-binding cassette (ABC) superfamily and address them to their plasma membrane to expel azole antifungals, which cancels out their action and allows the yeast to become multidrug resistant (MDR). In a way to understand this mechanism of defense, we describe the purification and characterization of Cdr1, the membrane ABC exporter mainly responsible for such phenotype in both species. Cdr1 proteins were functionally expressed in the baker yeast, tagged at their C-terminal end with either a His-tag for the </span><em>glabrata</em> version, <em>cg</em><span>Cdr1-His, or a green fluorescent protein (GFP) preceded by a proteolytic cleavage site for the </span><em>albicans</em> version, <em>ca</em><span>Cdr1-P-GFP. A membrane Cdr1-enriched fraction was then prepared to assay several detergents and stabilizers, probing their level of extraction and the ATPase activity of the proteins as a functional marker. Immobilized metal-affinity and size-exclusion chromatographies (IMAC, SEC) were then carried out to isolate homogenous samples. Overall, our data show that although topologically and phylogenetically close, both proteins display quite distinct behaviors during the extraction and purification steps, and qualify </span><em>cg</em>Cdr1 as a good candidate to characterize this type of proteins for developing future inhibitors of their azole antifungal efflux activity.</p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139055785","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-27DOI: 10.1016/S0300-9084(23)00318-8
{"title":"Inside front cover-EDB","authors":"","doi":"10.1016/S0300-9084(23)00318-8","DOIUrl":"https://doi.org/10.1016/S0300-9084(23)00318-8","url":null,"abstract":"","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0300908423003188/pdfft?md5=eca1935735368c0987b96f8b52490ca0&pid=1-s2.0-S0300908423003188-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139050118","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}
BiochimiePub Date : 2023-12-20DOI: 10.1016/j.biochi.2023.12.005
Valeriya V. Mikhaylova, Tatiana B. Eronina
{"title":"Effects of osmolytes under crowding conditions on the properties of muscle glycogen phosphorylase b","authors":"Valeriya V. Mikhaylova, Tatiana B. Eronina","doi":"10.1016/j.biochi.2023.12.005","DOIUrl":"10.1016/j.biochi.2023.12.005","url":null,"abstract":"<div><p><span><span>The study of the relationship between the activity and stability of enzymes under crowding conditions in the presence of osmolytes is important for understanding the functioning of a living cell. The effect of osmolytes (trehalose and betaine) on the secondary and </span>tertiary structure<span> and activity of muscle glycogen phosphorylase </span></span><em>b</em> (Ph<em>b</em><span><span><span>) under crowding conditions created by PEG 2000 and PEG 20000 was investigated using dynamic light scattering, </span>differential scanning calorimetry, circular dichroism spectroscopy, fluorimetry and </span>enzymatic activity assay. At 25 °C PEGs increased Ph</span><em>b</em><span> activity, but PEG 20000 to a greater extent. Wherein, PEG 20000 significantly destabilized its tertiary and secondary structure, in contrast to PEG 2000. Trehalose removed the effects of PEGs on Ph</span><em>b</em>, while betaine significantly reduced the activating effect of PEG 20000 without affecting the action of PEG 2000. Under heat stress at 48 °C, the protective effect of osmolytes under crowding conditions was more pronounced than at room temperature, and the Ph<em>b</em> activity in the presence of osmolytes was higher in these conditions than in diluted solutions. These results provide important insights into the complex mechanism, by which osmolytes affect the structure and activity of Ph<em>b</em> under crowding conditions.</p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138821184","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-19DOI: 10.1016/j.biochi.2023.12.003
Tanvir Bamra , Taj Shafi , Sushmita Das , Manjay Kumar , Pradeep Das
{"title":"Leishmania donovani mevalonate kinase regulates host actin for inducing phagocytosis","authors":"Tanvir Bamra , Taj Shafi , Sushmita Das , Manjay Kumar , Pradeep Das","doi":"10.1016/j.biochi.2023.12.003","DOIUrl":"10.1016/j.biochi.2023.12.003","url":null,"abstract":"<div><p>Despite the well-established role of macrophages in phagocytosing <span><em>Leishmania</em></span>, the contribution of the parasite to this process is not well understood. Present study provides insights into the mechanism underlying the MVK-induced entry of <em>L. donovani</em> and improve our knowledge of host-pathogen interactions. We have discussed <em>Mevalonate kinase (MVK)</em><span>-induced actin reorganization, modulation of signaling pathways and host cell functions. Our results show that LdMVK gains access to macrophage cytosol and induces actin assembly modulation through the activation of actin-related proteins: VASP, Src and ERM. We have also demonstrated that LdMVK induces Ca</span><sup>2+</sup> signaling and Akt pathway in macrophages, which are critical components of <em>Leishmania</em> survival and proliferation. Interestingly, we found that antibodies against LdMVK can kill <em>Leishmania</em>-infected macrophages in culture by forming extracellular traps, highlighting the potential of LdMVK in inhibiting parasite death. Overall, LdMVK is a virulent factor in <em>Leishmania</em><span><span><span> that mediates parasite internalization and host modulation by targeting host </span>proteins phosphorylation and </span>calcium homeostasis having significant implications in disease progression.</span></p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138741642","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-19DOI: 10.1016/j.biochi.2023.12.006
Margarita Beskrovnaia , Aleksei Agapov , Kristina Makasheva , Dmitry O. Zharkov , Daria Esyunina , Andrey Kulbachinskiy
{"title":"Sensing of DNA modifications by pAgo proteins in vitro","authors":"Margarita Beskrovnaia , Aleksei Agapov , Kristina Makasheva , Dmitry O. Zharkov , Daria Esyunina , Andrey Kulbachinskiy","doi":"10.1016/j.biochi.2023.12.006","DOIUrl":"10.1016/j.biochi.2023.12.006","url":null,"abstract":"<div><p><span><span>Many prokaryotic Argonaute<span><span> (pAgo) proteins act as programmable nucleases that use small guide </span>DNAs for recognition and cleavage of complementary target DNA. Recent studies suggested that pAgos participate in cell defense against invader DNA and may also be involved in other </span></span>genetic processes, including DNA replication and repair. The ability of pAgos to recognize specific targets potentially make them an invaluable tool for DNA manipulations. Here, we demonstrate that DNA-guided DNA-targeting pAgo nucleases from three bacterial species, DloAgo from </span><em>Dorea longicatena</em>, CbAgo from <span><em>Clostridium butyricum</em></span> and KmAgo from <em>Kurthia massiliensis</em><span>, can sense site-specific modifications in the target DNA, including 8-oxoguanine, thymine<span> glycol, ethenoadenine and pyrimidine dimers<span>. The effects of DNA modifications<span> on the activity of pAgos strongly depend on their positions relative to the site of cleavage and are comparable to or exceed the effects of guide-target mismatches at corresponding positions. For all tested pAgos, the strongest effects are observed when DNA lesions are located at the cleavage position. The results demonstrate that DNA cleavage by pAgos is strongly affected by DNA modifications, thus making possible their use as sensors of DNA damage.</span></span></span></span></p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138820548","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"New insights into the substrate specificity of cholesterol oxidases for more aware application","authors":"Michail Shapira , Alexandra Dobysh , Anastasia Liaudanskaya , Hanna Aucharova , Yaraslau Dzichenka , Volha Bokuts , Suzana Jovanović-Šanta , Aliaksey Yantsevich","doi":"10.1016/j.biochi.2023.12.004","DOIUrl":"10.1016/j.biochi.2023.12.004","url":null,"abstract":"<div><p><span>Cholesterol oxidases<span><span> (ChOxes) are enzymes that catalyze the </span>oxidation<span> of cholesterol to cholest-4-en-3-one. These enzymes find wide applications across various diagnostic and industrial settings. In addition, as a pathogenic factor of several bacteria, they have significant clinical implications. The current classification system for ChOxes is based on the type of bond connecting FAD to the apoenzyme, which does not adequately illustrate the enzymatic and structural characteristics of these proteins. In this study, we have adopted an integrative approach, combining evolutionary analysis, classic enzymatic techniques and computational approaches, to elucidate the distinct features of four various ChOxes from </span></span></span><span><em>Rhodococcus</em></span> sp. (RCO), <em>Cromobacterium</em> sp. (CCO), <span><em>Pseudomonas aeruginosa</em></span> (PCO) and <em>Burkhoderia cepacia</em><span><span> (BCO). Comparative and evolutionary analysis of substrate-binding domain (SBD) and FAD-binding domain (FBD) helped to reveal the origin of ChOxes. We discovered that all forms of ChOxes had a common ancestor and that the structural differences evolved later during divergence. Further examination of amino acid variations revealed SBD as a more variable compared to FBD independently of FAD coupling mechanism. Revealed differences in amino acid positions turned out to be critical in determining common for ChOxes properties and those that account for the individual differences in </span>substrate specificity. A novel look with the help of chemical descriptors on found distinct features were sufficient to attempt an alternative classification system aimed at application approach. While univocal characteristics necessary to establish such a system remain elusive, we were able to demonstrate the substrate and protein features that explain the differences in substrate profile.</span></p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138682498","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
BiochimiePub Date : 2023-12-16DOI: 10.1016/j.biochi.2023.12.002
Rebecca J. Webb , Catherine Rush , Lee Berger , Lee F. Skerratt , Alexandra A. Roberts
{"title":"Glutathione is required for growth and cadmium tolerance in the amphibian chytrid fungus, Batrachochytrium dendrobatidis","authors":"Rebecca J. Webb , Catherine Rush , Lee Berger , Lee F. Skerratt , Alexandra A. Roberts","doi":"10.1016/j.biochi.2023.12.002","DOIUrl":"10.1016/j.biochi.2023.12.002","url":null,"abstract":"<div><p><em>Batrachochytrium dendrobatidis</em> (<em>Bd</em>) is a lethal amphibian pathogen, partly due to its ability to evade the immune system of susceptible frog species. In many pathogenic fungi, the antioxidant glutathione is a virulence factor that helps neutralise oxidative stressors generated from host immune cells, as well as other environmental stressors such as heavy metals. The role of glutathione in stress tolerance in <em>Bd</em> has not been investigated. Here, we examine the changes in the glutathione pool after stress exposure and quantify the effect of glutathione depletion on cell growth and stress tolerance. Depletion of glutathione repressed growth and release of zoospores, suggesting that glutathione is essential for life cycle completion in <em>Bd.</em> Supplementation with <2 mM exogenous glutathione accelerated zoospore development, but concentrations >2 mM were strongly inhibitory to <em>Bd</em> cells. While hydrogen peroxide exposure lowered the total cellular glutathione levels by 42 %, glutathione depletion did not increase the sensitivity to hydrogen peroxide. Exposure to cadmium increased total cellular glutathione levels by 93 %. Glutathione-depleted cells were more sensitive to cadmium, and this effect was attenuated by glutathione supplementation, suggesting that glutathione plays an important role in cadmium tolerance. The effects of heat and salt were exacerbated by the addition of exogenous glutathione. The impact of glutathione levels on <em>Bd</em> stress sensitivity may help explain differences in host susceptibility to chytridiomycosis and may provide opportunities for synergistic therapeutics.</p></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":null,"pages":null},"PeriodicalIF":3.9,"publicationDate":"2023-12-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0300908423003164/pdfft?md5=4541ccba938f294987cfc8078f5551ec&pid=1-s2.0-S0300908423003164-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138682410","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}