{"refrec":{"BRefID":113509,"RR":"<b>Ken, C.-F.; Lin, C.-T.; Wen, Y.-D.; Wu, J.-L.</b> (2007). Replacement of buried cysteine from zebrafish Cu/Zn superoxide dismutase and enhancement of its stability via site-directed mutagenesis. <i>Mar. Biotechnol. 9(3)</i>: 335-342. <a href=\"https://dx.doi.org/10.1007/s10126-006-0143-y\" target=\"_blank\">https://dx.doi.org/10.1007/s10126-006-0143-y</a>","BEntID":107965,"PublicFlag":1,"CheckedFlag":1,"wosflag":1,"vabbflag":0,"RefStringPartII":". <i>Mar. Biotechnol. 9(3)</i>: 335-342. <a href=\"https://dx.doi.org/10.1007/s10126-006-0143-y\" target=\"_blank\">https://dx.doi.org/10.1007/s10126-006-0143-y</a>","DocTypID":8,"DocType":"Journal article","MarineFlag":0,"FreshFlag":0,"BrackishFlag":0,"TerrestrialFlag":0,"Authorstring":"Ken, C.-F.; Lin, C.-T.; Wen, Y.-D.; Wu, J.-L.","OrigTitleTranslFlag":0,"Authorstringtrunc":"Ken, C.-F. <i>et al.</i>","Englishabstract":"Zebrafish Cu/Zn-superoxide dismutase (ZSOD1) has one free cysteine (Cys-7) in a first β-strand with lower thermostability. We predicted the stability would be increased with single-point mutation at 70°C via the I-Mutant 2.0 server, and generated a mutant SOD with replacement of the free Cys to Ala (ZSODC7A) by site-directed mutagenesis. The mutant was expressed and purified from the <i>Escherichia coli</i> strain AD494(DE3)pLysS and the yield was 2 mg from 0.4 L of culture. The ZSODC7A was heated at 90°C. In a time-dependent assay, the time interval for 50% inactivation was 32 min, and its thermal inactivation rate constant <i>K</i><sub>d</sub> was 2 × 10<sup>−2</sup> min<sup>−1</sup>. The mutant was still activated in broad pH range (2.3–12), and had only a moderate effect under sodium dodecyl sulfate treatment. The calculated specific activity of the mutant was 3980 U/mg, twice that of wild-type ZSOD1. In addition, we soaked fish larva with equal enzyme units of either ZSOD1 or ZSODC7A for 2 h, and then stressed them with 100 ppm of paraquat to induce oxidative injury. The survival rate was significant.","AbstractOtherLang":null,"BibLvlCode":"AS","StandardTitle":"Replacement of buried cysteine from zebrafish Cu/Zn superoxide dismutase and enhancement of its stability via site-directed mutagenesis","OrigTitleLangCode":"en","OrigTitleLangCodeExtended":"eng","OrigTitleLangID":15,"DateLastModified":{"date":"2026-06-14 01:31:14.672383","timezone_type":1,"timezone":"+02:00"},"UserAccessRight":null,"UserAccID":null,"AuthorKeywords":"Cu/Zn superoxide dismutase; I-Mutant 2.0; oxidative stress; site-directmutagenesis; thermostability","OtherDescriptors":null,"Notes":null,"AnaPub":2007,"MonPub":null,"DateUpdate":"2020-10-23","DateCreate":"2007-08-29","SecASFANote":null,"ConfID":null,"PeerRev":1,"VlizCoreFlag":1,"WoScode":"WOS:000247207600004","VABBcode":null,"OpenAcc":0,"DOI":"10.1007/s10126-006-0143-y"},"refs":null,"anarec":{"AnaID":113509,"PubliDate":2007,"Pagination":"335-342","XtraPublOfAnaID":null,"ISBN":null,"Volume":"9","Issue":"3","BRefMon":null,"BRefMonRR":null,"BRefXtra":null,"BRefXtraRR":null,"SerBRefID":45426,"SerRR":"Marine Biotechnology. 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