کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
10537337 | 962714 | 2005 | 11 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Disulfide-linked dimers of human adrenaline synthesizing enzyme PNMT are catalytically active
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کلمات کلیدی
GSSGβMEAdoHcyS-adenosyl-l-homocysteinePNMTPAGEAdoMetGSHPEARMSDDTTSDSSECAUC - AUCS-adenosyl-L-methionine - S-adenosyl-L-metionineanalytical ultracentrifugation - ultracentrifugation تحلیلیβ-mercaptoethanol - β-merkaptoethanolEDTA - اتیلن دی آمین تترا استیک اسید polyacrylamide gel electrophoresis - الکتروفورز ژل پلی آکریل آمیدnative-PAGE - بومی PAGEDisulfide bond formation - تشکیل پیوند دی سولفیدProtein purification - خالصسازی یا تخلیص پروتئینdithiothreitol - دیتیوتریتولsodium dodecyl sulfate - سدیم دودسیل سولفاتCatecholamine synthesis - سنتز کاتچولامینDimerisation - فروپاشیCatalytic activity - فعالیت کاتالیستیPhenylethanolamine N-methyltransferase - فنیلتانولامین N-متیل ترانسفرازroot mean square deviation - میانگین انحراف مربع ریشهpolyethylene glycol - پلی اتیلن گلیکولPEG - پلیاتیلن گلیکول reduced glutathione - کاهش گلوتاتیونChromatography - کروماتوگرافیSize exclusion chromatography - کروماتوگرافی اندازهای طردیoxidized glutathione - گلوتاتیون اکسید شده
موضوعات مرتبط
مهندسی و علوم پایه
شیمی
شیمی آنالیزی یا شیمی تجزیه
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
The crystal structure of human phenylethanolamine N-methyltransferase (hPNMT) reveals a disulfide-linked dimer, despite the presence of reducing agent in the crystallisation conditions. By removing the reducing agent, hPNMT crystals grow more rapidly and at lower protein concentrations. However, it was unclear whether the disulfide bonds are only present in the crystal form or whether these affect enzyme activity. The solution oligomeric state of hPNMT was investigated using biochemical techniques and activity assays. We found that in the absence of reducing agent, hPNMT forms dimers in solution. Furthermore, the solution dimer of hPNMT incorporates disulfide bonds, since this form is sensitive to reducing agent. The C48A and C139A mutants of hPNMT, which are incapable of forming the disulfide bond observed in the crystal structure, have a decreased propensity to form dimer in solution. Those dimers that do form are also sensitive to reducing agent. Further, the C48A/C139A double mutant shows only monomeric behaviour. Both dimeric and monomeric hPNMT, as well as mutants have wildtype enzyme activity. These results show that a variety of disulfides, including those observed in the crystal structure, can form in solution. In addition, disulfide-linked dimers are as active as the monomeric enzyme indicating that the crystal structure of the protein is a valid target for inhibitor design.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1750, Issue 1, 15 June 2005, Pages 82-92
Journal: Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics - Volume 1750, Issue 1, 15 June 2005, Pages 82-92
نویسندگان
Christine L. Gee, Amanda Nourse, A-Yen Hsin, Qian Wu, Joel D. Tyndall, Gary L. Grunewald, Michael J. McLeish, Jennifer L. Martin,