Solid phase synthesis of a tetrapeptide labelled with Carbon-13. Preparation of L-[U-13C]lysyl-L-[1-13C]arginyl-L-[3-13C]asparyl-L-[1-13C]serine

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Abstract

Synthesis of a [13C] tetrapeptide L-[U-13C]Lysyl-L-[1-13C]Arginyl-L-[3-13C]Aspartyl-L-[1-13C] Serine (KRDS) on a p-alkoxybenzyl ester polystyrene-1% divinylbenzene resin support is described by using repetitive nonhydrolytic base cleavage of α-amino protective groups in Solid Phase Peptide Synthesis (SPPS). Protected [13C] amino acid (AA) used in this SPPS model were prepared with 9-Fluorenylmethyloxycarbonyl (Fmoc) protecting group and with different acid labile side chain protecting group over AA type as: Fmoc-Ser(tert-Bu)-OH, Fmoc-Asp(tert-Bu)-OH, Fmoc-Arg(Pmc)-OH and Fmoc-Lys(Fmoc)-OH. All the protected AA were coupled by N,N′-dicyclohexyl carbodiimide (DCC) procedure, followed by Fmoc group cleavage using 20% piperidine in N,N dimethylacetamide (DMA). Quantitative deblocking side-chain AA protection and removal of KRDS from the solid support was effected by treatment with 50% trifluoroacetic acid in methylene chloride in a one pot-procedure. Homogeneous free [13C] KRDS was obtained in 90% overall yield after HPLC purification. This synthesis schedule offered the advantage over present solid phase method which used acidolysis for repetitive α-amino group deblocking.

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