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Chemical Ligation of Glycopeptides

Talan, Rommel S.

Abstract Details

2010, Doctor of Philosophy, University of Toledo, Chemistry.

Advances in the chemical synthesis of homogeneous glycoproteins and glycopeptides have facilitated the structural studies of glycopeptides and glycoproteins, understanding the roles of glycans on protein function and development of glycopeptide-based therapeutics. Decarboxylative condensation and the reaction between thioacids and 2,4-dinitrobenzenesulfonamides are highly promising protocols for the chemoselective ligation of glycopeptide fragments that do not depend on thiol-capture methods and are relatively tolerant of steric crowding at the reaction centers. We have extended the synthetic scope of these ligation techniques to the synthesis of O-linked glycopeptides, as well as, the generation of N-linkage.

Investigations into decarboxylative condensation, as presented in Chapter 2, involved the synthesis of a series of protected and unprotected glycosyl dipeptides, which contained the α-keto acid moiety at the C-terminus, followed by their ligation to a series of O-tert-butyl protected N-hydroxylamino acids to afford O-tert-butyl-protected glycosyl tripeptides. The reactions were carried out under both anhydrous and aqueous conditions at neutral pH to produce glycopeptide products in yields ranging from 15% to 86% depending on the amino acids present at the ligation junction. The best yields were obtained when both the α-keto acid and the N-hydroxylamino acid contained medium-sized chains. In addition to the expected tripeptide product, 2,5-substituted oxazoles were isolated when O-tert-butyl protected N-hydroxylamines of glycine were employed in the reaction. The formation of the oxazole is believed to result from an intramolecular cyclization of the O-tert-butyl ester on a nitrilium ion intermediate followed by aromatization. A decarboxylative condensation between O18-labeled phenylpyruvic acid and N-hydroxyphenethylamine oxalate salt resulted in amide products lacking the O-18 label, providing further support for the nitrilium ion in the reaction pathway.

Integral to these efforts was the initial explorations of the solid-phase synthesis of the α-keto acid and the N-hydroxylamine which was laid out in Chapter 3. A select number of amino acids was loaded onto the Ellman “safety catch” 4-sulfamylbutyryl linker with loading efficiencies ranging from 26 to 80%. Activation of the amino acyl resin with iodoacetonitrile and subsequent displacement with alanine cyanoketophosphorane afforded the crude dipeptide cyanoketophosphorane product in very low yields. The requisite N,O-bis-Fmoc protected amino acids were obtained from the corresponding free hydroxylamines in excellent yields. The coupling of the bis-Fmoc alanine to the alanine-preloaded Wang-resin was incomplete due to severe steric hindrance which could not be resolved by using higher equivalents of the hydroxylamine, longer reaction times, and two coupling cycles. Cleavage of the dipeptide hydroxylamine from the resin was attended by premature loss of the Fmoc protecting groups.

The application of the amide bond-forming reaction between sulfonamides and thioacids to the synthesis of glycosyl amides and glycopeptides was demonstrated in Chapter 4. β-N-glucosyl asparagine was synthesized from the corresponding N-glucosylsulfonamide and β-thioaspartic acid in good yields at mild conditions and short reaction times. A series of N-peptidyl and N-glycopeptidyl-2,4-dinitrobenzenesulfonamides consisting of short MUC 1 sequences was successfully assembled on the solid phase following Fmoc/tBu strategy with yields from 40% to 85%. Likewise, a series of Boc- and Fmoc-protected amino trityl thioesters were prepared from the corresponding amino acids in good to excellent yields. The analogous preparation of peptide thioesters suffered from low yields and epimerization. The reaction of the side-chain unprotected thioacids derived from the trityl thioesters with alanine sulfonamide proceeded chemoselectively and efficiently, especially at hindered junctions. Similarly, the ligation of histidine thioacid with N-glycopentapeptidylsulfonamide bearing unprotected peptide functionalities and protected glycan afforded the desired product in good yield (71%).

Steven Sucheck, PhD (Advisor)
Max Funk, PhD (Committee Member)
Mark Mason, PhD (Committee Member)
James Slama, PhD (Committee Member)
396 p.

Recommended Citations

Citations

  • Talan, R. S. (2010). Chemical Ligation of Glycopeptides [Doctoral dissertation, University of Toledo]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=toledo1278958947

    APA Style (7th edition)

  • Talan, Rommel. Chemical Ligation of Glycopeptides. 2010. University of Toledo, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=toledo1278958947.

    MLA Style (8th edition)

  • Talan, Rommel. "Chemical Ligation of Glycopeptides." Doctoral dissertation, University of Toledo, 2010. http://rave.ohiolink.edu/etdc/view?acc_num=toledo1278958947

    Chicago Manual of Style (17th edition)