Convergent solid-phase peptide synthesis

Lloyd‐Williams, Paul; Alberício, Fernando; Giralt, Ernest

doi:10.1016/s0040-4020(01)81800-7

Cited by 212 publications

(107 citation statements)

References 402 publications

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“…The repetition of the MUC-1 20-mer peptide is an attractive model with which to study the efficiency of the convergent approach for the solid-phase synthesis of large peptides and small proteins (24). The method is based on the principle that protected peptide fragments corresponding to the entire protein sequence can be condensed sequentially on a suitable solid support.…”

Section: Resultsmentioning

confidence: 99%

Preparation of MUC-1 Oligomers Using an Improved Convergent Solid-phase Peptide Synthesis

Krambovitis¹,

Hatzidakis²,

Barlos³

1998

Journal of Biological Chemistry

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The sequentially repeating nature of the core mucin polypeptide chain MUC-1 on the surface of malignant cells makes it an excellent target for cancer immunotherapy. We describe a reliable and efficient method of synthesizing oligomers, up to five tandem repeats and oligomer heterotope derivatives with a 15-amino acid epitope from tetanus toxin using an improved convergent solid-phase peptide synthesis. The different oligomers were easily distinguishable by reverse-phase high pressure liquid chromatography, but they were poorly fixed and migrated with the same migration rate, irrespective of size, in electrophoretic studies. In contrast, the oligomer heterotopes exhibited size-dependent electrophoretic behavior but in high pressure liquid chromatography chromatograms the different heterotopes were eluted simultaneously in two peaks representing the L-and D-enantiomers of the derivatives. The oligomer heterotopes were recognized as antigens in Western blotting with a murine monoclonal antibody against the epitope APDTR. In enzyme immunoassay studies with the same antibody an increasing reactivity was observed against the larger oligomers and confirmed by inhibition assays as the MUC-1 pentamer was the most efficient inhibitor. These results support the suggestion that the pentamer attains a structure closer to the native conformation and is more immunogenic. In conclusion, large composite peptides can be reliably synthesized with the convergent solid-phase peptide strategy offering an attractive option to vaccine designing and development.

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Section: Resultsmentioning

confidence: 99%

Preparation of MUC-1 Oligomers Using an Improved Convergent Solid-phase Peptide Synthesis

Krambovitis¹,

Hatzidakis²,

Barlos³

1998

Journal of Biological Chemistry

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“…We next evaluated the feasibility of a more challenging proposition, namely, the possibility of an iterative chemoselective solidphase fragment coupling (SPFC) via isonitrile-mediated amidation. Due to the high levels of convergence, SPFC is, in principle, an attractive strategy for the synthesis of large peptides and/or peptides with "difficult" sequences (34)(35)(36)(37)(38)(39)(40). Moreover, SPFC may offer considerable advantages in allowing for interim purification, which should simplify obtaining homogeneous end product.…”

Section: % (mentioning

confidence: 99%

“…However, ligations in the SPPS mode have often been plagued by limitations. Protected peptide fragments, which are used in traditional SPFC, are difficult to purify by HPLC due to their poor solubility in aqueous solvents (34,35). To solve this problem, it would be ideal if minimally protected peptide fragments could be used in the ligation step.…”

Section: % (mentioning

confidence: 99%

Solid-phase peptide synthesis and solid-phase fragment coupling mediated by isonitriles

Wang

2013

Proc. Natl. Acad. Sci. U.S.A.

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The synthesis of polypeptides on solid phase via mediation by isonitriles is described. The acyl donor is a thioacid, which presumably reacts with the isonitrile to generate a thio-formimidate carboxylate mixed anhydride intermediate. Applications of this chemistry to reiterative solid-phase peptide synthesis as well as solid-phase fragment coupling are described.A mide bond formations are arguably among the most important constructions in organic chemistry (1, 2). The centrality of the amide linkage, as found in polypeptides and proteins, in the maintenance of life hardly needs restatement. Numerous strategies, resulting in a vast array of protocols to synthesize biologically active polypeptides and proteins, have been demonstrated (3, 4). Central to reiterative polypeptide bond formations was the discovery and remarkable development of solid-phase peptide synthesis (SPPS) (5, 6). The extraordinary impact of SPPS in fostering enhanced access to homogeneous polypeptides is clear to everyone in the field.As we have described elsewhere, by classical, mechanistic reasoning, we were led to conjecture about some hithertounexplored possibilities relevant to the chemistry of isonitriles (7)(8)(9)(10)(11)(12)(13)(14). It was anticipated that isonitriles might be able to mediate the acylation of amines, thus giving rise to amides (15). Early experiments focused on free carboxylic acids as the acylating agents. As our studies progressed, it was found that the combination of thioacids, amines, and isonitriles leads to the efficient formation of amide bonds under stoichiometric or nearstoichiometric conditions (7-13, 16, 17). Although there remain unresolved issues of detail and nuance, the governing mechanism for amide formation under these conditions involves reaction of the thioacid, 1, with an isonitrile, 2, to generate a thio-formimidate carboxylate mixed anhydride (thio-FCMA), 3, which is intercepted by the "acyl-accepting" amine to generate amide, 5, and thioformamide, 6 (Fig. 1). The efficiency of the amidation was further improved through the use of hydroxybenzotriazole (HOBt) (18), which could well give rise to HOBt ester 7, although this pathway has not been mechanistically proven.The potentialities of the isonitrile-mediated amidation method were foreshadowed via its application to the synthesis of cyclosporine (19). The power of the method was particularly well demonstrated in the context of our recent total synthesis of oxytocin (OT) (20), wherein isonitrile mediation was used in each of the peptide bond constructions, leading to the synthesis of the hormone in high yield and excellent purity. This nonapeptide is involved in a range of biological functions including parturition and lactation (21,22). Signaling of OT to its receptor (OTR) is apparently an important factor in quality maintenance of various CNS functions (23). The ability to synthesize such modestly sized, but bio-impactful peptides in both native (wild-type) form, and as strategically modified variants, is one of the current missions of our labo...

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“…Such impurities are very difficult to identify and to separate from the required product. These problems can be overcome by the convergent peptide synthesis (CPS) strategy [1][2][3][4] where, instead of amino acids, fully characterized and homogenous protected segments are applied. To enable the simple SPPS of protected peptide segments with free the C-terminal carboxyl group, a variety of resins and linkers were developed.…”

Section: Introductionmentioning

confidence: 99%

9-Fluorenylmethyloxycarbonyl/tbutyl-based convergent protein synthesis

Barlos

Gatos

1999

Biopolymers

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Besides linear solid phase peptide synthesis, segment condensation in solution and chemical ligation, convergent peptide synthesis (CPS) was developed in order to enable the efficient preparation of complex peptides and small proteins. According to this synthetic strategy, solid phase synthesized and suitably protected peptide fragments corresponding to the entire peptide/protein‐sequence are condensed on a solid support or in solution, to the target protein. This review summarizes CPS performed utilizing the mild 9‐fluorenylmethyloxycarbonyl/tbutyloxycarbonyl‐based protecting scheme for the amino acids. © 1999 John Wiley & Sons, Inc. Biopoly 51: 266–278, 1999

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Convergent solid-phase peptide synthesis

Cited by 212 publications

References 402 publications

Preparation of MUC-1 Oligomers Using an Improved Convergent Solid-phase Peptide Synthesis

Preparation of MUC-1 Oligomers Using an Improved Convergent Solid-phase Peptide Synthesis

Solid-phase peptide synthesis and solid-phase fragment coupling mediated by isonitriles

9-Fluorenylmethyloxycarbonyl/tbutyl-based convergent protein synthesis

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