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Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010What Could Be the Role of Quinacrine inCreutzfeldt-Jakob Disease Treatment?Zbigniew Zawada 1,2 , Jaroslav Šebestík 1 , Martin Šafařík 1 ,Anna Krejčiříková 3 , Anna Březinová 1 , Jan Hlaváček 1 ,Ivan Stibor 1 , Karel Holada 3 , and Petr Bouř 11 Institute of Organic Chemistry and Biochemistry, Academy of Sciences CR, Flemingovonám. 2, Prague 6, 166 10, Czech Republic; 2 Institute of Chemical Technology, Technická 5,166 28 Prague 6, Czech Republic; 3 First Faculty of Medicine, Charles University inPrague, Kateřinská 32, Prague 2, Czech RepublicIntroductionA conversion from normal cellular form of prion protein (PrP C ) to a toxic one (PrP Sc ) issuspected to cause prion diseases [1] such as Kuru, Creutzfeldt-Jakob disease, mad cowdisease, etc. This transformation can be prevented, at least in vitro, by quinacrine [2].Quinacrine can react with primary amines [3] and thiols [4] to afford acridine analogs(Figure 1), what was utilized in our laboratory during the study of acridinylation reaction offree thiol groups of prion protein.Results and DiscussionSince quinacrine prevents plaque formation in vitro [2] and it reacts with thiols, wesuggested that the main action of quinacrine in prevention of plaque formation in cell is itsreaction with thio groups of PrP cysteine residues. To find out the affinity of quinacrinetowards PrP we started with affinity study of short fragments of PrP containing cysteineresidue (Table 1).ClQuinacrineNRSHClNOONHS RNFig. 1. Acridinylation reaction of primary thiol.We were able to obtain two types of acridinylated peptides:a) peptides where acridine moiety is attached just to the sulfur atom of cysteine (1, 2,3, 4, 5, 6, 7, 8), andb) peptides where the acridine moiety is attached not only to the cysteine’s sulfuratom, but also to the N-terminus amino group (1, 2, 4, 5, 6).In the case of longer peptides 7 and 8 we were able to obtain just traces ofbisacridinylated product, where both acridine moieties are attached to sulfur atom ofcysteine residues. In these cases we were not able to observe any N-acridinylation. Themost probable reason is poor solubility of both peptides under physiological conditions(phosphate buffer, pH=7.4, 37°C).To prove that it is the N-terminus amino group that is acridinylated, and not histidine’simidazole ring, we prepared a set of three peptides 1, 2 and 3 with almost the samesequence. Whereas we were not able to obtain any bisacridinylated N-acetylated peptide 3,we were able to obtain bisacridinylated peptides 1 and 2. It means that histidine is notnecessary for the second acridinylation, but the N-terminus amino group is essential. Wesupported this hypothesis also by NMR spectra.We found that it is possible to lead the reaction course toward almost pure cysteineS-acridinylation by lowering the pH value of reaction mixture, by using subequimolar amountof quinacrine and by shortening the reaction time. Bisacridinylated peptides can be obtained asa main product when huge excess of quinacrine is used in higher pH for longer reaction time.84