Total synthesis of bovine pancreatic trypsin inhibitor and the protein diastereomer [Gly37D‐Ala]BPTI using Boc chemistry solid phase peptide synthesis

Abstract

AbstractBovine pancreatic trypsin inhibitor (BPTI) is a well‐studied model for investigation of protein folding and stability. Here, we report the synthesis and characterization of wild‐type BPTI and a diastereomeric protein analogue [Gly37D‐Ala]BPTI. Each 58‐residue polypeptide chain was made by native chemical ligation of two peptide segments, BPTI[1‐29]‐αthioester and the appropriate version of the Cys30‐58 BPTI peptide segment. Boc chemistry in situ neutralization solid phase synthesis was used to prepare the peptide segment reactants. The resulting full‐length polypeptide chains were folded in a cysteine/cystine redox buffer to give synthetic protein molecules containing three disulfide bonds. The diastereomeric analogue [Gly37D‐Ala]BPTI folded as efficiently as the native protein. Synthetic proteins were characterized by analytical LCMS and by natural‐abundance 1H‐15N HSQC NMR fingerprinting. These results illustrate the power of Boc chemistry peptide synthesis and its utility for the total chemical synthesis of protein molecules.