{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,8]],"date-time":"2026-10-08T16:02:47Z","timestamp":1791475367661,"version":"4.3.4"},"reference-count":46,"publisher":"Springer Science and Business Media LLC","issue":"5","license":[{"start":{"date-parts":[[2025,7,18]],"date-time":"2025-07-18T00:00:00Z","timestamp":1752796800000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"},{"start":{"date-parts":[[2025,7,18]],"date-time":"2025-07-18T00:00:00Z","timestamp":1752796800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/501100004477","name":"Stellenbosch University","doi-asserted-by":"crossref","id":[{"id":"10.13039\/501100004477","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":["link.springer.com"],"crossmark-restriction":false},"short-container-title":["Protein J"],"published-print":{"date-parts":[[2025,10]]},"abstract":"<jats:title>Abstract<\/jats:title>\n          <jats:p>\n            <jats:italic>Plasmodium falciparum<\/jats:italic>, the parasite responsible for the most severe form of malaria, encodes three small heat shock proteins (sHsps), PfHsp20a, PfHsp20b, and PfHsp20c, each containing a conserved \u03b1-crystallin domain (ACD). These class I sHsps are hypothesized to play critical roles in proteostasis under stress, yet their specific functions have remained poorly defined. In this study, all three sHsps were recombinantly expressed and purified for structural and functional characterization. Circular dichroism and thermal shift assays revealed distinct conformational properties, with PfHsp20a exhibiting the highest thermal and chemical stability. Functional assays using malate dehydrogenase and citrate synthase confirmed that all three isoforms possess autonomous chaperone activity, although with varying efficiency. Notably, the plant-derived flavonoid quercetin disrupted both the structure and function of the sHsps in a concentration-dependent manner, with PfHsp20c being the most sensitive. Quercetin also inhibited the growth of <jats:italic>P. falciparum<\/jats:italic> Nf54 and Dd2 strains in vitro with IC<jats:sub>50<\/jats:sub> values of 5.4\u00a0\u03bcM and 7.8\u00a0\u03bcM, respectively. These results provide the first direct evidence of independent chaperone activity in <jats:italic>P. falciparum<\/jats:italic> sHsps and highlight their vulnerability to small molecule inhibition. This establishes their potential as novel drug targets for antimalarial intervention.<\/jats:p>","DOI":"10.1007\/s10930-025-10281-w","type":"journal-article","created":{"date-parts":[[2025,7,18]],"date-time":"2025-07-18T16:56:02Z","timestamp":1752857762000},"page":"580-597","update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Comparative Characterization of Plasmodium falciparum Small Heat Shock Proteins and Their Inhibition by Quercetin (3,3\u2032,4\u2032,5,7-Pentahydroxyflavone)"],"prefix":"10.1007","volume":"44","author":[{"given":"Francisca Magum","family":"Timothy","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tawanda","family":"Zininga","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,7,18]]},"reference":[{"key":"10281_CR1","unstructured":"World Health Organization. World Malaria Report 2023; available at https:\/\/www.who.int\/teams\/global-malaria-programme\/reports\/world-malaria-report-2023"},{"key":"10281_CR2","doi-asserted-by":"publisher","first-page":"1216192","DOI":"10.3389\/fmolb.2023.1216192","volume":"10","author":"SY Almaazmi","year":"2023","unstructured":"Almaazmi SY, Kaur RP, Singh H, Blatch GL (2023) The Plasmodium falciparum exported J domain proteins fine-tune human and malarial Hsp70s: pathological exploitation of proteostasis machinery. Front Mol Biosci 10:1216192","journal-title":"Front Mol Biosci"},{"key":"10281_CR3","doi-asserted-by":"publisher","first-page":"323","DOI":"10.1146\/annurev-biochem-060208-092442","volume":"82","author":"YE Kim","year":"2013","unstructured":"Kim YE, Hipp MS, Bracher A, Hayer-Hartl M, Hartl FU (2013) Molecular chaperone functions in protein folding and proteostasis. Annu Rev Biochem 82:323\u2013355","journal-title":"Annu Rev Biochem"},{"issue":"6","key":"10281_CR4","doi-asserted-by":"publisher","first-page":"2121","DOI":"10.1074\/jbc.REV118.002809","volume":"294","author":"M Haslbeck","year":"2019","unstructured":"Haslbeck M, Weinkauf S, Buchner J (2019) Small Heat shock proteins: simplicity meets complexity. J Biol Chem 294(6):2121","journal-title":"J Biol Chem"},{"key":"10281_CR5","doi-asserted-by":"publisher","first-page":"7","DOI":"10.1007\/s12192-018-0948-4","volume":"24","author":"HH Kampinga","year":"2019","unstructured":"Kampinga HH et al (2019) Function, evolution, and structure of J-domain proteins. Cell Stress Chaperones 24:7\u201315","journal-title":"Cell Stress Chaperones"},{"issue":"1","key":"10281_CR6","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2021.167157","volume":"434","author":"K Reinle","year":"2022","unstructured":"Reinle K, Mogk A, Bukau B (2022) The diverse functions of small heat shock proteins in the proteostasis network. J Mol Biol 434(1):167157","journal-title":"J Mol Biol"},{"issue":"2","key":"10281_CR7","doi-asserted-by":"publisher","first-page":"295","DOI":"10.1007\/s12192-019-00979-z","volume":"24","author":"S Carra","year":"2019","unstructured":"Carra S et al (2019) Small heat shock proteins: multifaceted proteins with important implications for life. Cell Stress Chaperones 24(2):295\u2013308","journal-title":"Cell Stress Chaperones"},{"key":"10281_CR8","doi-asserted-by":"publisher","first-page":"89","DOI":"10.1146\/annurev-micro-020518-115515","volume":"73","author":"A Mogk","year":"2019","unstructured":"Mogk A, Ruger-Herreros C, Bukau B (2019) Cellular functions and mechanisms of action of small heat shock proteins. Annu Rev Microbiol 73:89\u2013110","journal-title":"Annu Rev Microbiol"},{"issue":"1","key":"10281_CR9","doi-asserted-by":"publisher","first-page":"1","DOI":"10.1186\/s12870-022-03587-9","volume":"22","author":"Y Zhou","year":"2022","unstructured":"Zhou Y et al (2022) Small HSPs play an important role in crosstalk between HSF-HSP and ROS pathways in heat stress response. BMC Plant Biol 22(1):1\u201316","journal-title":"BMC Plant Biol"},{"issue":"7","key":"10281_CR10","doi-asserted-by":"publisher","first-page":"1537","DOI":"10.1016\/j.jmb.2015.02.002","volume":"427","author":"M Haslbeck","year":"2015","unstructured":"Haslbeck M, Vierling E (2015) A first line of stress defense: small heat shock proteins and their function in protein homeostasis. J Mol Biol 427(7):1537\u20131548","journal-title":"J Mol Biol"},{"issue":"7","key":"10281_CR11","doi-asserted-by":"publisher","first-page":"2112","DOI":"10.3390\/ijms19072112","volume":"19","author":"VM Shatov","year":"2018","unstructured":"Shatov VM et al (2018) Role of the arginine in the conserved N-terminal domain RLFDQxFG motif of human small heat shock proteins. Int J Mol Sci 19(7):2112","journal-title":"Int J Mol Sci"},{"key":"10281_CR12","doi-asserted-by":"publisher","first-page":"842149","DOI":"10.3389\/fmolb.2022.842149","volume":"9","author":"B Tedesco","year":"2022","unstructured":"Tedesco B et al (2022) Insights on human small heat shock proteins and their alterations in diseases. Front Mol Biosci 9:842149","journal-title":"Front Mol Biosci"},{"issue":"1","key":"10281_CR13","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1038\/s41467-020-20314-w","volume":"12","author":"C Yu","year":"2021","unstructured":"Yu C et al (2021) Structural basis of substrate recognition and thermal protection by a small heat shock protein. Nat Commun 12(1):1\u201311","journal-title":"Nat Commun"},{"issue":"4","key":"10281_CR14","doi-asserted-by":"publisher","first-page":"457","DOI":"10.1007\/s12192-009-0155-4","volume":"15","author":"G Kapp\u00e9","year":"2010","unstructured":"Kapp\u00e9 G, Boelens WC, de Jong WW (2010) Why proteins without an \u03b1-crystallin domain should not be included in the human small heat shock protein family HSPB. Cell Stress Chaperones 15(4):457\u2013461","journal-title":"Cell Stress Chaperones"},{"issue":"4","key":"10281_CR15","doi-asserted-by":"publisher","first-page":"581","DOI":"10.1007\/s12192-020-01093-1","volume":"25","author":"WC Boelens","year":"2020","unstructured":"Boelens WC (2020) Structural aspects of the human small heat shock proteins related to their functional activities. Cell Stress Chaperones 25(4):581","journal-title":"Cell Stress Chaperones"},{"issue":"10","key":"10281_CR16","doi-asserted-by":"publisher","first-page":"a034025","DOI":"10.1101\/cshperspect.a034025","volume":"11","author":"MK Janowska","year":"2019","unstructured":"Janowska MK, Baughman HER, Woods CN, Klevit RE (2019) Mechanisms of small heat shock proteins. Cold Spring Harb Perspect Biol 11(10):a034025","journal-title":"Cold Spring Harb Perspect Biol"},{"issue":"2\u20133","key":"10281_CR17","doi-asserted-by":"publisher","first-page":"108","DOI":"10.1016\/j.plaphy.2010.01.001","volume":"48","author":"AO Tiroli-Cepeda","year":"2010","unstructured":"Tiroli-Cepeda AO, Ramos CHI (2010) Heat causes oligomeric disassembly and increases the chaperone activity of small heat shock proteins from sugarcane. Plant Physiol Biochem 48(2\u20133):108\u2013116","journal-title":"Plant Physiol Biochem"},{"issue":"8","key":"10281_CR18","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0105892","volume":"9","author":"M Heirbaut","year":"2014","unstructured":"Heirbaut M, Beelen S, Strelkov SV, Weeks SD (2014) Dissecting the functional role of the N-terminal domain of the human small heat shock protein HSPB6. PLoS ONE 9(8):e105892","journal-title":"PLoS ONE"},{"key":"10281_CR19","doi-asserted-by":"publisher","first-page":"1322780","DOI":"10.3389\/fceld.2023.1322780","volume":"2","author":"FM Timothy","year":"2023","unstructured":"Timothy FM, Zininga T (2023) Small heat shock proteins as modulators of cell death in Plasmodium falciparum parasites and its human host. Front Cell Death 2:1322780","journal-title":"Front Cell Death"},{"issue":"52","key":"10281_CR20","doi-asserted-by":"publisher","first-page":"39943","DOI":"10.1074\/jbc.M607677200","volume":"281","author":"E Basha","year":"2006","unstructured":"Basha E, Friedrich KL, Vierling E (2006) The N-terminal arm of small heat shock proteins is important for both chaperone activity and substrate specificity. J Biol Chem 281(52):39943\u201339952","journal-title":"J Biol Chem"},{"issue":"1","key":"10281_CR21","doi-asserted-by":"publisher","first-page":"241","DOI":"10.1007\/s00289-022-04091-8","volume":"80","author":"M Azeem","year":"2023","unstructured":"Azeem M et al (2023) An insight into anticancer, antioxidant, antimicrobial, antidiabetic and anti-inflammatory effects of quercetin: a review. Polym Bull 80(1):241\u2013262","journal-title":"Polym Bull"},{"issue":"1","key":"10281_CR22","first-page":"7632408","volume":"2024","author":"H Memariani","year":"2024","unstructured":"Memariani H, Memariani M, Ghasemian A (2024) Quercetin as a promising antiprotozoan phytochemical: current knowledge and future research avenues. Biomed Res Int 2024(1):7632408","journal-title":"Biomed Res Int"},{"issue":"13","key":"10281_CR23","doi-asserted-by":"publisher","first-page":"6776","DOI":"10.3390\/ijms22136776","volume":"22","author":"A Burger","year":"2021","unstructured":"Burger A et al (2021) Characterization of an atypical Trypanosoma brucei Hsp70 demonstrates its cytosolic-nuclear localization and modulation by quercetin and methylene blue. Int J Mol Sci 22(13):6776","journal-title":"Int J Mol Sci"},{"issue":"6","key":"10281_CR24","doi-asserted-by":"publisher","DOI":"10.1371\/journal.pone.0129445","volume":"10","author":"T Zininga","year":"2015","unstructured":"Zininga T et al (2015) Overexpression, purification and characterisation of the Plasmodium falciparum Hsp70-z (PfHsp70-z) Protein. PLoS ONE 10(6):e0129445","journal-title":"PLoS ONE"},{"issue":"4","key":"10281_CR25","doi-asserted-by":"publisher","first-page":"409","DOI":"10.1007\/s12551-016-0218-6","volume":"8","author":"J Stetefeld","year":"2016","unstructured":"Stetefeld J, McKenna SA, Patel TR (2016) Dynamic light scattering: a practical guide and applications in biomedical sciences. Biophys Rev 8(4):409","journal-title":"Biophys Rev"},{"issue":"3","key":"10281_CR26","doi-asserted-by":"publisher","first-page":"554","DOI":"10.1111\/febs.16312","volume":"290","author":"JAJ Housmans","year":"2023","unstructured":"Housmans JAJ et al (2023) A guide to studying protein aggregation. FEBS J 290(3):554\u2013583","journal-title":"FEBS J"},{"issue":"5","key":"10281_CR27","doi-asserted-by":"publisher","first-page":"957","DOI":"10.1002\/prot.26779","volume":"93","author":"FL Muzenda","year":"2025","unstructured":"Muzenda FL et al (2025) Characterization and inhibition of the chaperone function of Plasmodium falciparum glucose-regulated protein 94 kDa (PfGrp94). Proteins 93(5):957\u2013971","journal-title":"Proteins"},{"issue":"4","key":"10281_CR28","doi-asserted-by":"publisher","first-page":"102688","DOI":"10.1016\/j.xpro.2023.102688","volume":"4","author":"T Wu","year":"2023","unstructured":"Wu T et al (2023) Protocol for performing and optimizing differential scanning fluorimetry experiments. STAR Protoc. 4(4):102688","journal-title":"STAR Protoc."},{"issue":"1","key":"10281_CR29","doi-asserted-by":"publisher","first-page":"19","DOI":"10.1002\/pro.3703","volume":"29","author":"C Sun","year":"2020","unstructured":"Sun C, Li Y, Yates EA, Fernig DG (2020) SimpleDSFviewer: a tool to analyze and view differential scanning fluorimetry data. Protein Sci 29(1):19","journal-title":"Protein Sci"},{"issue":"3","key":"10281_CR30","doi-asserted-by":"publisher","first-page":"499","DOI":"10.1007\/s12192-016-0678-4","volume":"21","author":"T Zininga","year":"2016","unstructured":"Zininga T et al (2016) Plasmodium falciparum Hsp70-z Exhibits independent chaperone activity and interacts with Hsp70-1 in a nucleotide-dependent fashion. Cell Stress Chaperones 21(3):499\u2013513","journal-title":"Cell Stress Chaperones"},{"issue":"2","key":"10281_CR31","doi-asserted-by":"publisher","first-page":"379","DOI":"10.1042\/BJ20031958","volume":"381","author":"TK Chowdary","year":"2004","unstructured":"Chowdary TK et al (2004) Mammalian Hsp22 is a heat-inducible small heat-shock protein with chaperone-like activity. Biochem J 381(2):379\u2013387","journal-title":"Biochem J"},{"issue":"1","key":"10281_CR32","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2021.167201","volume":"434","author":"M Vendruscolo","year":"2022","unstructured":"Vendruscolo M, Fuxreiter M (2022) Sequence determinants of the aggregation of proteins within condensates. J Mol Biol 434(1):167201","journal-title":"J Mol Biol"},{"issue":"16","key":"10281_CR33","doi-asserted-by":"publisher","first-page":"2072","DOI":"10.1002\/1873-3468.14449","volume":"596","author":"A Horvath","year":"2022","unstructured":"Horvath A et al (2022) Are casein micelles extracellular condensates formed by liquid-liquid phase separation? FEBS Lett 596(16):2072\u20132085","journal-title":"FEBS Lett"},{"issue":"3","key":"10281_CR34","doi-asserted-by":"publisher","first-page":"295","DOI":"10.1016\/j.ijpddr.2017.07.001","volume":"7","author":"S Duffy","year":"2017","unstructured":"Duffy S, Avery VM (2017) Plasmodium falciparum in vitro continuous culture conditions: a comparison of drug susceptibility. Int J Parasitol Drugs Drug Resist 7(3):295\u2013302","journal-title":"Int J Parasitol Drugs Drug Resist"},{"issue":"1","key":"10281_CR35","doi-asserted-by":"publisher","first-page":"37","DOI":"10.3390\/ijms25010037","volume":"25","author":"O Adamczyk","year":"2024","unstructured":"Adamczyk O et al (2024) Bovine serum albumin as a platform for designing biologically active nanocarriers. Int J Mol Sci 25(1):37","journal-title":"Int J Mol Sci"},{"issue":"52","key":"10281_CR36","doi-asserted-by":"publisher","first-page":"33254","DOI":"10.1073\/pnas.2007670117","volume":"117","author":"M Hardenberg","year":"2020","unstructured":"Hardenberg M et al (2020) Widespread occurrence of the droplet state of proteins in the human proteome. Proc Natl Acad Sci USA 117(52):33254\u201333262","journal-title":"Proc Natl Acad Sci USA"},{"key":"10281_CR37","doi-asserted-by":"publisher","DOI":"10.3389\/fmolb.2022.947203","volume":"9","author":"T Muthelo","year":"2022","unstructured":"Muthelo T et al (2022) Inhibition of Plasmodium falciparum Hsp70-Hop partnership by 2-phenylthynesulfonamide. Front Mol Biosci 9:947203","journal-title":"Front Mol Biosci"},{"issue":"2","key":"10281_CR38","doi-asserted-by":"publisher","first-page":"78","DOI":"10.4161\/cam.20101","volume":"6","author":"GN Montagna","year":"2012","unstructured":"Montagna GN, Matuschewski K, Buscaglia CA (2012) Small heat shock proteins in cellular adhesion and migration: evidence from Plasmodium genetics. Cell Adhes Migr 6(2):78\u201384","journal-title":"Cell Adhes Migr"},{"issue":"4","key":"10281_CR39","doi-asserted-by":"publisher","first-page":"2410","DOI":"10.1074\/jbc.M111.302109","volume":"287","author":"GN Montagna","year":"2012","unstructured":"Montagna GN et al (2012) Critical role for heat shock protein 20 (HSP20) in migration of malarial sporozoites. J Biol Chem 287(4):2410\u20132422","journal-title":"J Biol Chem"},{"key":"10281_CR40","doi-asserted-by":"publisher","first-page":"1063","DOI":"10.1016\/j.tibs.2024.08.003","volume":"40","author":"C Peters","year":"2024","unstructured":"Peters C, Haslbeck M, Buchner J (2024) Catchers of folding gone awry: a tale of small heat shock proteins. Trends Biochem Sci 40:1063","journal-title":"Trends Biochem Sci"},{"issue":"2","key":"10281_CR41","doi-asserted-by":"publisher","first-page":"144","DOI":"10.1016\/j.molbiopara.2006.03.011","volume":"148","author":"C Mehlin","year":"2006","unstructured":"Mehlin C et al (2006) Heterologous expression of proteins from Plasmodium falciparum: results from 1000 genes. Mol Biochem Parasitol 148(2):144\u2013160","journal-title":"Mol Biochem Parasitol"},{"issue":"2\u20133","key":"10281_CR42","doi-asserted-by":"publisher","first-page":"79","DOI":"10.1016\/j.bpc.2009.09.003","volume":"145","author":"AS Kazakov","year":"2009","unstructured":"Kazakov AS et al (2009) Thermally induced structural changes of intrinsically disordered small heat shock protein Hsp22. Biophys Chem 145(2\u20133):79\u201385","journal-title":"Biophys Chem"},{"issue":"5","key":"10281_CR43","doi-asserted-by":"publisher","first-page":"3512","DOI":"10.1021\/acs.analchem.9b05712","volume":"92","author":"E Vuorinen","year":"2020","unstructured":"Vuorinen E et al (2020) Sensitive label-free thermal stability assay for protein denaturation. Anal Chem 92(5):3512\u20133516","journal-title":"Anal Chem"},{"issue":"2","key":"10281_CR44","doi-asserted-by":"publisher","first-page":"672","DOI":"10.1074\/jbc.M116.760413","volume":"292","author":"EV Mymrikov","year":"2017","unstructured":"Mymrikov EV et al (2017) The Chaperone activity and substrate spectrum of human small heat shock proteins. J Biol Chem 292(2):672\u2013684","journal-title":"J Biol Chem"},{"issue":"35","key":"10281_CR45","doi-asserted-by":"publisher","first-page":"9362","DOI":"10.1021\/acs.jpcb.6b06214","volume":"120","author":"A Precupas","year":"2016","unstructured":"Precupas A, Sandu R, Popa VT (2016) Quercetin influence on thermal denaturation of bovine serum albumin. J Phys Chem B 120(35):9362\u20139375","journal-title":"J Phys Chem B"},{"issue":"2","key":"10281_CR46","doi-asserted-by":"publisher","first-page":"90","DOI":"10.1038\/nchem.1243","volume":"4","author":"GR Bickerton","year":"2012","unstructured":"Bickerton GR et al (2012) Quantifying the chemical beauty of drugs. Nat Chem 4(2):90\u201398","journal-title":"Nat Chem"}],"container-title":["The Protein Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s10930-025-10281-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/article\/10.1007\/s10930-025-10281-w\/fulltext.html","content-type":"text\/html","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/link.springer.com\/content\/pdf\/10.1007\/s10930-025-10281-w.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,9,24]],"date-time":"2025-09-24T13:05:15Z","timestamp":1758719115000},"score":1,"resource":{"primary":{"URL":"https:\/\/link.springer.com\/10.1007\/s10930-025-10281-w"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,7,18]]},"references-count":46,"journal-issue":{"issue":"5","published-print":{"date-parts":[[2025,10]]}},"alternative-id":["10281"],"URL":"https:\/\/doi.org\/10.1007\/s10930-025-10281-w","relation":{},"ISSN":["1572-3887","1875-8355"],"issn-type":[{"value":"1572-3887","type":"print"},{"value":"1875-8355","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,7,18]]},"assertion":[{"value":"30 June 2025","order":1,"name":"accepted","label":"Accepted","group":{"name":"ArticleHistory","label":"Article History"}},{"value":"18 July 2025","order":2,"name":"first_online","label":"First Online","group":{"name":"ArticleHistory","label":"Article History"}},{"order":1,"name":"Ethics","group":{"name":"EthicsHeading","label":"Declarations"}},{"value":"The authors declare no competing interests.","order":2,"name":"Ethics","group":{"name":"EthicsHeading","label":"Conflict of interest"}}]}}