Proceedings book download - 5Z.com

Proceedings of the 31 st European Peptide SymposiumMichal Lebl, Morten Meldal, Knud J. Jensen, Thomas Hoeg-Jensen (Editors)European Peptide Society, 2010Oxyfold: A New Solid Supported Reagent for the Simple andEffective Formation of Disulfide Bond in PeptidesL. Ronga 1 , P. Verdié 2 , M. Cristau 2 , M. Amblard 2 , S. Cantel 2 ,C. Enjalbal 2 , K. Puget 1 , G. Subra 2 , and J. Martinez 21 Genepep, St Clément de Rivière, 34980, France, 2 Institut des Biomolécules MaxMousseron (IBMM), Montpellier, 34093, FranceIntroductionThe importance of disulfide bridges in peptide conformation and activity make theirformation a fundamental step of peptide synthesis. Formation of disulfide bond is probablyone of the most challenging steps to achieve regarding the formation of unwantedby-products and oligomerization. In order to minimize the latter phenomenon, disulfidebridge cyclizations are performed under high diluted conditions which require timeconsuming removal of solvent at the end of the reaction. Dimethyl sulfoxide (DMSO) isone of the oxidizing reactants most commonly used [1]. One of its major drawbacks is itselimination from the reaction medium, which requires evaporation under strong vacuum orrepeated lyophilizations. Furthermore, the dimethyl sulfide generated during the reaction isvolatile and toxic. On the other hand, supported reactants are particularly advantageous forpromoting intramolecular reactions. In fact, they are known to cause a phenomenon of“pseudodilution” which makes it possible to minimize oligomerization and to use muchsmaller amounts of solvents [2]. Here we present the synthesis and the use of novel solidsupported oxidation reactants for disulfide bond formation. This family of supportedreagents consists in a series of oxidized methionines grafted onto a solid support. Wedemonstrate the efficiency and easiness of these supported reagents for the formation ofdisulfide bridges in peptides.Results and DiscussionThree different solid supported methionine sulfoxides were synthesized by anchoring aFmoc-protected Met on different matrices: Amino PEGA, Amino PEG-PS andaminomethyl polystyrene. After Fmoc deprotection and acetylation, methionine oxidationwas performed by treatment of the resin with H 2 O 2 . The ability of these supportedmethionine sulfoxides to promote disulfide bond formation of crustacean cardioactivepeptide CCAP (PFCNAFTGC-NH 2 ) [3] was explored at pH=6.1 (ammonium acetatebuffer) and pH=7.5 (phosphate buffer). Under the same conditions, the CCAP oxidationpromoted by Ellman’s reagent prepared on amino PEGA resin and 10% DMSO solutionwas investigated. Our results (Figure 1) show that Oxyfold is more effective than DMSOand supported Ellman’s reagent. Among the compared resins, amino PEGA resin gives thebest results of CCAP oxidation. Moreover, pH=7.5 seems the ideal pH for Oxyfoldpromotedoxidation.pH 6.1pH 7.5100100% oxydized CCAP80604020% oxydized CCAP8060402000 5 10 15 20 25 30time (h)00 5 10 15 20 25 30time (h)Fig. 1. CCAP oxidation: ■ 10% DMSO,▲ Ac-Met[O]-PEGA,● Ac-Met[O]-PS, ♦ Ellman’sreagent-PEGA,× Ac-Met[O]-PEG-PS.182