Title

Synthesis of Polyalanine Peptides Containing a Lysine Residue via Solid-Phase Peptide

Format

Oral Presentation

Abstract/Artist Statement

The synthesis of a polyalanine peptide with a single lysine residue has been achieved using the solid-phase peptide synthesis (SPPS) method. Our SPPS method involves coupling the 9- fluorenylmethyloxycarbonyl (Fmoc) protected amino acid residues sequentially to a solid support using the coupling reagents HOBt (1-hydroxybenzotriazole) and DIC (diisoproplycarbodiimide). The solid support consists of small insoluble beads with linkers that attach the C-terminus of the peptide covalently by an ester linkage. This allows for the ease of purification by eliminating impurities through washing with dichloromethane and N,N- dimethylformamide (DMF) while the peptides are immobilized by an insoluble solid support. Successive couplings are performed by removal of the base- labile Fmoc group using diethylamine and followed by addition of the next desired Fmoc protected residue. The deprotection and coupling procedures are repeated until the desired peptide length is reached. Once completed, the N-terminus is protected by an acetyl group using acetic anhydride and the peptide is cleaved from the solid support with the cleavage cocktail Reagent R, mainly composed of trifluoroacetic acid (TFA). The lysine side chain is protected by an acid labile t-butoxycarbonyl (t-Boc) protecting group and is cleaved off concurrently with the cleavage of the solid support. Purification is accomplished either by chloroform extraction or precipitation in cold diethyl ether. Analysis of the peptide and sequence determination is performed by using a triple quadruple mass spectrometer coupled with electrospray ionization. Successful synthesis was confirmed by obtaining the expected ionized fragments. Several peptides have been successfully synthesized, including the pentapeptide Ac- KAAAA.

Location

University of the Pacific, Classroom Building

Start Date

5-5-2007 9:00 AM

End Date

5-5-2007 12:30 PM

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May 5th, 9:00 AM May 5th, 12:30 PM

Synthesis of Polyalanine Peptides Containing a Lysine Residue via Solid-Phase Peptide

University of the Pacific, Classroom Building

The synthesis of a polyalanine peptide with a single lysine residue has been achieved using the solid-phase peptide synthesis (SPPS) method. Our SPPS method involves coupling the 9- fluorenylmethyloxycarbonyl (Fmoc) protected amino acid residues sequentially to a solid support using the coupling reagents HOBt (1-hydroxybenzotriazole) and DIC (diisoproplycarbodiimide). The solid support consists of small insoluble beads with linkers that attach the C-terminus of the peptide covalently by an ester linkage. This allows for the ease of purification by eliminating impurities through washing with dichloromethane and N,N- dimethylformamide (DMF) while the peptides are immobilized by an insoluble solid support. Successive couplings are performed by removal of the base- labile Fmoc group using diethylamine and followed by addition of the next desired Fmoc protected residue. The deprotection and coupling procedures are repeated until the desired peptide length is reached. Once completed, the N-terminus is protected by an acetyl group using acetic anhydride and the peptide is cleaved from the solid support with the cleavage cocktail Reagent R, mainly composed of trifluoroacetic acid (TFA). The lysine side chain is protected by an acid labile t-butoxycarbonyl (t-Boc) protecting group and is cleaved off concurrently with the cleavage of the solid support. Purification is accomplished either by chloroform extraction or precipitation in cold diethyl ether. Analysis of the peptide and sequence determination is performed by using a triple quadruple mass spectrometer coupled with electrospray ionization. Successful synthesis was confirmed by obtaining the expected ionized fragments. Several peptides have been successfully synthesized, including the pentapeptide Ac- KAAAA.