{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,18]],"date-time":"2025-10-18T10:34:54Z","timestamp":1760783694019},"reference-count":43,"publisher":"Wiley","issue":"9","license":[{"start":{"date-parts":[[2013,8,1]],"date-time":"2013-08-01T00:00:00Z","timestamp":1375315200000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Protein Science"],"published-print":{"date-parts":[[2013,9]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>The bifunctional enzyme thymidylate synthase\u2013dihydrofolate reductase (TS\u2013DHFR) plays an essential role in DNA synthesis and is unique to several species of pathogenic protozoans, including the parasite <jats:italic>Toxoplasma gondii<\/jats:italic>. Infection by <jats:italic>T. gondii<\/jats:italic> causes the prevalent disease toxoplasmosis, for which TS\u2013DHFR is a major therapeutic target. Here, we design peptides that target the dimer interface between the TS domains of bifunctional <jats:italic>T. gondii<\/jats:italic> TS\u2013DHFR by mimicking \u03b2\u2010strands at the interface, revealing a previously unknown allosteric target. The current study shows that these \u03b2\u2010strand mimetic peptides bind to the apo\u2010enzyme in a species\u2010selective manner to inhibit both the TS and distal DHFR. Fluorescence spectroscopy was used to monitor conformational switching of the TS domain and demonstrate that these peptides induce a conformational change in the enzyme. Using structure\u2010guided mutagenesis, nonconserved residues in the linker between TS and DHFR were identified that play a key role in domain\u2013domain communication and in peptide inhibition of the DHFR domain. These studies validate allosteric inhibition of apo\u2010TS, specifically at the TS\u2013TS interface, as a potential target for novel, species\u2010specific therapeutics for treating <jats:italic>T. gondii<\/jats:italic> parasitic infections and overcoming drug resistance.<\/jats:p>","DOI":"10.1002\/pro.2300","type":"journal-article","created":{"date-parts":[[2013,6,27]],"date-time":"2013-06-27T17:40:34Z","timestamp":1372354834000},"page":"1161-1173","source":"Crossref","is-referenced-by-count":8,"title":["Selective peptide inhibitors of bifunctional thymidylate synthase\u2010dihydrofolate reductase from <i>Toxoplasma gondii<\/i> provide insights into domain\u2013domain communication and allosteric regulation"],"prefix":"10.1002","volume":"22","author":[{"given":"Mark","family":"J. Landau","sequence":"first","affiliation":[{"name":"Department of Pharmacology Yale University School of Medicine 333 Cedar Street New Haven Connecticut 06510"},{"name":"Department of Molecular Biophysics and Biochemistry Yale University School of Medicine 333 Cedar Street New Haven Connecticut 06510"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hitesh","family":"Sharma","sequence":"additional","affiliation":[{"name":"Department of Pharmacology Yale University School of Medicine 333 Cedar Street New Haven Connecticut 06510"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Karen S.","family":"Anderson","sequence":"additional","affiliation":[{"name":"Department of Pharmacology Yale University School of Medicine 333 Cedar Street New Haven Connecticut 06510"},{"name":"Department of Molecular Biophysics and Biochemistry Yale University School of Medicine 333 Cedar Street New Haven Connecticut 06510"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2013,8]]},"reference":[{"key":"e_1_2_8_2_1","doi-asserted-by":"publisher","DOI":"10.1098\/rsif.2006.0115"},{"key":"e_1_2_8_3_1","doi-asserted-by":"publisher","DOI":"10.1002\/jcp.20356"},{"key":"e_1_2_8_4_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.3099389"},{"key":"e_1_2_8_5_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00050a007"},{"key":"e_1_2_8_6_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi991610i"},{"key":"e_1_2_8_7_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi952953t"},{"key":"e_1_2_8_8_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M206523200"},{"key":"e_1_2_8_9_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.5560010611"},{"key":"e_1_2_8_10_1","doi-asserted-by":"publisher","DOI":"10.1107\/S1744309111029009"},{"key":"e_1_2_8_11_1","doi-asserted-by":"publisher","DOI":"10.1038\/nsb0394-186"},{"key":"e_1_2_8_12_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M310328200"},{"key":"e_1_2_8_13_1","doi-asserted-by":"publisher","DOI":"10.1107\/S090744490901230X"},{"key":"e_1_2_8_14_1","doi-asserted-by":"publisher","DOI":"10.1038\/nsb921"},{"key":"e_1_2_8_15_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00387a052"},{"key":"e_1_2_8_16_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.exppara.2004.03.002"},{"key":"e_1_2_8_17_1","doi-asserted-by":"publisher","DOI":"10.1128\/AAC.48.6.2116-2123.2004"},{"key":"e_1_2_8_18_1","doi-asserted-by":"publisher","DOI":"10.1001\/archopht.1960.01840010142016"},{"key":"e_1_2_8_19_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi701624u"},{"key":"e_1_2_8_20_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bmcl.2008.11.054"},{"key":"e_1_2_8_21_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi952923q"},{"key":"e_1_2_8_22_1","doi-asserted-by":"publisher","DOI":"10.1042\/BJ20081247"},{"key":"e_1_2_8_23_1","doi-asserted-by":"crossref","first-page":"6269","DOI":"10.1016\/S0021-9258(18)53249-X","article-title":"Primary structure of the dihydrofolate reductase\u2010thymidylate synthase gene from Toxoplasma gondii","volume":"268","author":"Roos DS","year":"1993","journal-title":"J Biol Chem"},{"key":"e_1_2_8_24_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.1104829108"},{"key":"e_1_2_8_25_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi9720989"},{"key":"e_1_2_8_26_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi400576t"},{"key":"e_1_2_8_27_1","doi-asserted-by":"publisher","DOI":"10.1107\/S0907444910044732"},{"key":"e_1_2_8_28_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi7019386"},{"key":"e_1_2_8_29_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.171"},{"key":"e_1_2_8_30_1","doi-asserted-by":"publisher","DOI":"10.1002\/pro.539"},{"key":"e_1_2_8_31_1","doi-asserted-by":"publisher","DOI":"10.1016\/j.bbapap.2003.09.005"},{"key":"e_1_2_8_32_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0167-4838(02)00289-3"},{"key":"e_1_2_8_33_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M009493200"},{"key":"e_1_2_8_34_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi061309j"},{"key":"e_1_2_8_35_1","doi-asserted-by":"publisher","DOI":"10.1007\/s00239-011-9433-8"},{"key":"e_1_2_8_36_1","doi-asserted-by":"publisher","DOI":"10.1021\/jm300850v"},{"key":"e_1_2_8_37_1","doi-asserted-by":"publisher","DOI":"10.1098\/rsob.120120"},{"key":"e_1_2_8_38_1","doi-asserted-by":"publisher","DOI":"10.1038\/188231a0"},{"key":"e_1_2_8_39_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi700531r"},{"key":"e_1_2_8_40_1","volume-title":"The PyMol Molecular Graphics System","author":"DeLano WL","year":"2002"},{"key":"e_1_2_8_41_1","doi-asserted-by":"publisher","DOI":"10.1093\/bioinformatics\/btm076"},{"key":"e_1_2_8_42_1","doi-asserted-by":"publisher","DOI":"10.1074\/jbc.M212689200"},{"key":"e_1_2_8_43_1","doi-asserted-by":"publisher","DOI":"10.1006\/prep.1997.0750"},{"key":"e_1_2_8_44_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00410a022"}],"container-title":["Protein Science"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1002%2Fpro.2300","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/api.wiley.com\/onlinelibrary\/tdm\/v1\/articles\/10.1002%2Fpro.2300","content-type":"unspecified","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/pdf\/10.1002\/pro.2300","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2023,9,15]],"date-time":"2023-09-15T11:42:36Z","timestamp":1694778156000},"score":1,"resource":{"primary":{"URL":"https:\/\/onlinelibrary.wiley.com\/doi\/10.1002\/pro.2300"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2013,8]]},"references-count":43,"journal-issue":{"issue":"9","published-print":{"date-parts":[[2013,9]]}},"alternative-id":["10.1002\/pro.2300"],"URL":"https:\/\/doi.org\/10.1002\/pro.2300","archive":["Portico"],"relation":{},"ISSN":["0961-8368","1469-896X"],"issn-type":[{"value":"0961-8368","type":"print"},{"value":"1469-896X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2013,8]]}}}