Chemical protein synthesis by solid phase ligation of unprotected peptide segments

In this paper we describe "solid phase chemical ligation" (SPCL), the appli cation of the principles of polymer-supported organic synthesis to the cons truction of large polypeptide chains for the total chemical synthesis of pr oteins. In this method, each building block used is an unprotected peptide segment of 20 or more amino acids. These are consecutively reacted by chemi cal ligation, the chemoselective reaction of the unprotected peptide segmen ts from aqueous solution, to make the polymer-supported target polypeptide. In a final step, the assembled full-length target polypeptide is released from the aqueous-compatible polymer support. Here we report chemistries for the attachment of the first segment to a polymer support, and for the asse mbly of the target polypeptide chain starting from the polymer-bound peptid e segment. In this solid phase protein synthesis method, large target polyp eptide chains can be built efficiently and rapidly by SPCL and, after relea se from the polymer support, folded to give functional protein molecules. S everal examples of the application of SPCL are given: model peptides consis ting of 27 and 68 amino acids, and polypeptides corresponding to the protei ns C5a (74 amino acids) and MIF(IIS amino acids), were each made in good yi eld and purity from the consecutive solid phase ligation of peptide segment s. In addition, we report the total synthesis by SPCL of the enzyme "human group V secretory phospholipase A(2)" (GV-PLA(2)), which comprises a polype ptide of 118 amino acids containing 6 disulfide bonds. As demonstrated by t hese examples, SPCL is an important extension of our capabilities for total chemical protein synthesis.