Tandem Ligation of Multipartite Peptides with Cell-Permeable Activity

• Published in: Journal of the American Chemical Society Vol. 125; no. 1; pp. 73 - 82
• Main Authors: Eom, Khee Dong, Miao, Zhenwei, Yang, Jin-Long, Tam, James P
• Format: Journal Article
• Language: English
• Published: WASHINGTON American Chemical Society 08.01.2003; Amer Chemical Soc
• Subjects: Aminoacids, peptides. Hormones. Neuropeptides; Analytical, structural and metabolic biochemistry; Biological and medical sciences; Biological Transport; Carrier Proteins - chemical synthesis; Carrier Proteins - chemistry; Carrier Proteins - pharmacokinetics; Cell Membrane Permeability; Chemistry; Chemistry, Multidisciplinary; Chromatography, High Pressure Liquid; Cysteine - chemistry; Cysteine - metabolism; Epitopes, T-Lymphocyte - chemistry; Epitopes, T-Lymphocyte - metabolism; Fundamental and applied biological sciences. Psychology; HeLa Cells; Humans; Microscopy, Confocal; Peptides - chemical synthesis; Peptides - chemistry; Peptides - pharmacokinetics; Physical Sciences; Proline - analogs & derivatives; Proline - metabolism; Proteins; Science & Technology; Structure-Activity Relationship; ANTIBACTERIAL PEPTIDES; PROLINE-RICH; AMINO-ACID-SEQUENCE; NATIVE CHEMICAL LIGATION; UNPROTECTED PEPTIDES; ANTIMICROBIAL PEPTIDE; ORTHOGONAL LIGATION; SYNTHETIC PEPTIDE; T-CELL; PROTEIN TRANSDUCTION; Synthesis; Peptides; Cell permeability; Tandem reaction
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja020529r

To prepare multipartite peptides with several functional cargoes including a cell-permeable sequence or transportant for intracellular delivery, tandem ligation of peptides is a convenient convergent approach with the fewest synthetic steps. It links three or four unprotected segments forming two or more regiospecific bonds consecutively without a deprotection step. This paper describes a tandem ligation strategy to prepare multipartite peptides with normal and branched architectures carrying a novel transportant peptide that is rich in arginine and proline to permit their cargoes to be translocated across membranes to affect their biological functions in cytoplasm. Our strategy consists of three ligation methods specific for amino terminal cysteine (Cys), serine/threonine (Ser/Thr), and Nα-chloroacetylated amine to afford Xaa-Cys, Xaa-OPro (oxaproline) and Xaa-ψGly (pseudoglycine) at the ligation sites, respectively. Assembly of single-chain peptides from three different segments was achieved by the tandem Cys/OPro ligation to form two amide bonds, an Xaa-Cys and then an Xaa-OPro. Assembly of two- and three-chain peptides with branched architectures from four different segments was accomplished by tandem Cys/ψGly/OPro ligation. These NT-specific tandem ligation strategies were successful in generating cell-permeable multipartite peptides with one-, two-, and three-chain architectures, ranging in size from 52 to 75 residues and without the need of a protection or deprotection step. In addition, our results show that there is considerable flexibility in architectural design to obtain cell-permeable multipartite peptides containing a transportant sequence.