{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,6,30]],"date-time":"2026-06-30T08:22:54Z","timestamp":1782807774858,"version":"3.54.5"},"reference-count":27,"publisher":"Wiley","issue":"5","license":[{"start":{"date-parts":[[2022,4,9]],"date-time":"2022-04-09T00:00:00Z","timestamp":1649462400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01_AI45999"],"award-info":[{"award-number":["R01_AI45999"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["U19AI089681"],"award-info":[{"award-number":["U19AI089681"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["onlinelibrary.wiley.com"],"crossmark-restriction":true},"short-container-title":["Protein Science"],"published-print":{"date-parts":[[2022,5]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>The <jats:italic>Plasmodium<\/jats:italic> ookinete uses chitinase activity to penetrate the acellular, chitin\u2010containing peritrophic matrix to invade the mosquito vector. <jats:italic>Plasmodium<\/jats:italic> ookinetes from different parasite clades secrete two structurally distinct forms of chitinase, one, a short form lacking a C\u2010terminal putative chitin\u2010binding domain (CBD), the other, a long form with both proenzyme and C\u2010terminal putative chitin\u2010binding domains. Here, we structurally and functionally characterize the three cysteines in the short chitinase of the human\u2010infecting malaria parasite, <jats:italic>P. falciparum<\/jats:italic> testing the hypothesis that one unpaired cysteine would not contribute to chitinase\u2010specific enzymatic activity which would identify this residue as potentially involved in intermolecular disulfide bonding and heteromultimeric invasion complex formation as previously described. To test this hypothesis, we produced and characterized recombinant wild\u2010type and cysteine\u2010mutation PfCHT1 proteins in <jats:italic>E. coli<\/jats:italic> and used biophysical and enzymatic approaches to examine their enzymatic activities and chitin\u2010binding affinities. The cysteine\u2010203 PfCHT1 mutation had no effect on chitinolytic and chitin\u2010binding functions. The cysteine\u2010220 and cysteine\u2010230 mutants were enzymatically inactive and did not bind to chitin. The artificial intelligence\u2010based protein prediction algorithm, AlphaFold, correctly identified the involvement of cys\u2010220 and cys\u2010230 in the intramolecular disulfide linkages key to maintaining properly folded chitinase structural integrity. AlphaFold predicted that cys\u2010203 cysteine is surface exposed and thus involved in intermolecular protein\u2013protein interaction. Production of the cys\u2010to\u2010ser 203 PfCHT1 mutant facilitated recombinant protein production. Future cellular and biochemical studies are needed to further understand details of <jats:italic>Plasmodium<\/jats:italic> ookinete mosquito midgut invasion.<\/jats:p>","DOI":"10.1002\/pro.4289","type":"journal-article","created":{"date-parts":[[2022,4,9]],"date-time":"2022-04-09T14:47:14Z","timestamp":1649515634000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Structure\u2013function analysis of cysteine residues in the <i>plasmodium falciparum<\/i> chitinase, <scp>PfCHT1<\/scp>"],"prefix":"10.1002","volume":"31","author":[{"given":"Hargobinder","family":"Kaur","sequence":"first","affiliation":[{"name":"Section of Infectious Diseases, Department of Internal Medicine Yale School of Medicine  New Haven Connecticut USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Laine","family":"Garber","sequence":"additional","affiliation":[{"name":"Section of Infectious Diseases, Department of Internal Medicine Yale School of Medicine  New Haven Connecticut USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"James W.","family":"Murphy","sequence":"additional","affiliation":[{"name":"Department of Pharmacology Yale University School of Medicine  New Haven Connecticut USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8344-2004","authenticated-orcid":false,"given":"Joseph M.","family":"Vinetz","sequence":"additional","affiliation":[{"name":"Section of Infectious Diseases, Department of Internal Medicine Yale School of Medicine  New Haven Connecticut USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2022,4,9]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.1016\/S1471-4922(01)01918-3"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.275.14.10331"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.70.3.1581-1590.2002"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1086\/432552"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1016\/0014-4894(91)90132-G"},{"key":"e_1_2_7_7_1","doi-asserted-by":"publisher","DOI":"10.1128\/IAI.69.6.4041-4047.2001"},{"key":"e_1_2_7_8_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.90.9.4266"},{"key":"e_1_2_7_9_1","doi-asserted-by":"publisher","DOI":"10.1111\/cmi.12413"},{"key":"e_1_2_7_10_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.96.24.14061"},{"key":"e_1_2_7_11_1","doi-asserted-by":"crossref","first-page":"e0031421","DOI":"10.1128\/IAI.00314-21","article-title":"Evolutionary insights into the microneme\u2010secreted, chitinase\u2010containing high molecular weight protein complexes involved in plasmodium invasion of the mosquito midgut","volume":"90","author":"Kaur H","year":"2021","journal-title":"Infect Immun"},{"key":"e_1_2_7_12_1","doi-asserted-by":"publisher","DOI":"10.3389\/fcimb.2020.615343"},{"key":"e_1_2_7_13_1","doi-asserted-by":"publisher","DOI":"10.1038\/s41586-021-03819-2"},{"key":"e_1_2_7_14_1","doi-asserted-by":"publisher","DOI":"10.1038\/d41586-020-03348-4"},{"key":"e_1_2_7_15_1","first-page":"1144","article-title":"The game has changed","volume":"370","author":"Service RF","year":"2020","journal-title":"AI Triumphs Protein Fold Sci"},{"key":"e_1_2_7_16_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0166-6851(03)00140-3"},{"key":"e_1_2_7_17_1","first-page":"223","article-title":"Chitinases of human parasites and their implications as antiparasitic targets","volume":"87","author":"Shahabuddin 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