Head-to-tail cyclic peptides with a wide range of ring sizes have been discovered in various organisms including bacteria, fungi, plants and animals. Many of them exhibit remarkable biological activities with high potency. Daptomycin, a cyclic lipodepsipeptide isolated from soil bacteria Stretomyces roseoporus, is the first natural product antibiotic launched in a generation. Daptomycin has potent bactericidal activity against otherwise antibiotic-resistant Gram-positive pathogens including methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE) and vancomycin-resistant S. aureus. Daptomycin contains a 31-membered ring made up of 10 amino acids and a linear 3-amino acid side chain modified with an n-decanoyl lipid at the N-terminus. The complex structure of daptomycin, the presence of two non-proteinogenic amino acids (kynurenine and 3-methyl glutamic acid) and the macrolactamization of a 31-membered ring render daptomycin a challenging target for total synthesis. We recently developed a chemoselective serine/threonine ligation (STL) allowing peptide ligation at Ser/Thr site using side chain unprotected segments. We have successfully applied STL intramolecularly in the key cyclization step for the synthesis of daptomycin molecule, which allows us to finally achieve the first total synthesis of daptomycin. With this technique in hand, the chemical synthesis of daptomycin analogues, which are difficult to obtain otherwise becomes possible and is now ongoing. This would allow for the search for optimized analogues. We further investigated if STL could be applied for the synthesis of cyclic tetrapeptides, which are extremely difficult to be synthesized in the absence of turninducing components due to their rigid structural framework. To our delight, a series of cyclic tetrapeptides without any turn-inducing component, like Gly, Pro or Damino acids have been successfully synthesized by intramolecular STL. The synthesis of cyclic tetrapeptides with drug-like scaffold would be useful for the therapeutic development. We also applied intramolecular STL to successfully synthesize some natural cyclic peptides with different ring sizes, including anticancer stylopeptide 1, phakellistatin 4 and anti-inflammatory cyclosquamosin D. / published_or_final_version / Chemistry / Doctoral / Doctor of Philosophy

D.Sc. (Chemistry) / Please refer to full text to view abstract

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The syntheses of s-triazolo[3,4-a]phthalazines (15, 3-R-TAP) by reaction of hydralazine (12, 1-hydrazinophthalazine) with N-protected amino acids and dipeptides under homogeneous (solution) and heterogeneous ("solid-phase") conditions are reported. Transition metal complexes containing the TAP ligand were prepared and their spectral properties investigated. The use of metal-ions and a cation-exchange resin (H+ form) were considered for the mild hydrolysis of side-chain amide bonds in TAP derivatives. The objective of these studies was to determine the feasibility of reacting the carboxyl groups in amino acids with hydralazine to afford the TAP derivatives as a method for peptide sequencing from the C-terminal residue. Hydralazine reacts with carboxylic acids to form an amide intermediate which undergoes ring closure with elimination of water to form the s-triazolo[3,4-a]phthalazine derivative. To promote the initial binding of hydralazine to the acid, coupling reagents were used to activate the carboxylate group towards nucleophilic attack. N-Ethyl-5-phenylisoxazolium-3'-sulfonate (17, NEPIS), 1-ethoxy -carbonyl-2-ethoxy-l,2-dihydroquinoline (27, EEDQ), various phosphorus compounds, carbodiimides, and chloroformates were carboxyl activating agents used to synthesize TAP derivatives. In solution studies, the carbodiimides (EDC, 16 and DCC, Si), NEPIS (17), and a combination of triphenylphosphite with imidazole are the most successful procedures for TAP synthesis In solid-phase studies, the best procedures for activating immobilized amino acids, are with isobuty'l chloroformate, NEPIS (l?), and DCC (51). Transition metal complexes were synthesized with the general formula: [M(3-H-TAP) (H20)6_n](C104)m (n = 4, m = 2, M = Co, Ni, Cu; n = 2, m = 2, M = Ni; n = 6, m = 3, M = Co). The infrared and visible spectra of these complexes are reported. [Co(.trien)C3-(N-Ac-gly)-TAP)J(C104)2 was also prepared and under acidic conditions, no hydrolysis of the side-chain amide bond was observed. There was also no significant hydrolysis of the side-chain with free ?+ 2+ 3-(N-Ac-gly)-TAP in the presence of Co and Cu under acidic conditions, or when it was eluted through a cation-exchange (H+ form) column. The decomposition of hydralazine in non-aqueous media was investigated and a major product of the decomposition was identified as diphthalazinylhydrazine {82). The implication of our studies is that the modification of amino acids with hydralazine is not yet a viable method for C-terminal peptide sequencing. Improvements are required for improving the yields of the coupled product, and the lack of a mild and selective method for hydrolyzing the C-terminal peptide bond limits the method at present to determination of the C-terminal residue only. / Science, Faculty of / Chemistry, Department of / Graduate

published_or_final_version / abstract / Chemistry / Doctoral / Doctor of Philosophy

A number of α-melanotropin analogues have been prepared. Insight towards the development and understanding of the functional roles of each of the amino acid residues important for high melanotropic potency in a number of biological systems was studied. The melanotropin analogue Ac- α-MSH₄₋₁₂-NH₂ contains all of the structural requirements necessary for obtaining full biological potency on the lizard (Anolis carolinensis) melanophore and S-91 mouse melanoma. On the frog (Rana pipiens) melanophore, the melanotropin analogue Ac-[Nle⁴]-α-MSH₄₋₁₃-NH₂ possesses full melanotropic potency relative to the native hormone. The smallest melanotropin analogue capable of eliciting any biological response was Ac-α-MSH₆₋₉-NH₂ (Ac-His-Phe-Arg-Trp-NH₂) on all biological systems studied. The low biological activity found in previous studies for smaller melanotropin analogues may have been due to a trace contamination of a potent melanotropin. The importance of each of the thirteen amino acid residues of α-melanotropin in contributing to the melanotropic actions in a number of biological systems is discussed. We were unable to confirm the reported presence of a second independent active sequence in α-melanotropin. The structural requirements for prolongation of biological activity (following removal of exogenous hormone from the assay medium) and enzyme stability have been studied. Analysis suggests species-dependent differences in the structural relationships of the amino acid residues in the 4, 7 and 11 positions for prolonged melanotropic response. Analogues prepared with D-phenylalanine in place of its L-enantiomer in the seventh positions of α-melanotropin analogues generally results in an increased resistance to enzymatic degradation towards rat brain homogenate, rat serum, and to the purified enzymes, trypsin and chymotrypsin. Some of these analogues have been shown to be useful probes for the understanding of melanotropic actions in a number of biological systems. Two α-melanotropin analogues have been prepared which possess partial agonism on the mouse melanoma adenylate cyclase assay. A number of these analogues should prove useful in future studies directed towards a more detailed study of melanotropin receptor systems in a large variety of biological systems. The first known synthesis of melanin concentrating hormone (MCH) is reported. The overall synthetic yield of this cyclic heptadecapeptide was 14%. The chemical, physical and biological properties of synthetic MCH and naturally occurring telost MCH were in agreement. MCH was found to be a full agonist in stimulating melanosome dispersion in both the frog and lizard bioassay. Therefore MCH can stimulate melanosome dispersion or contraction depending upon the bioassay studied. Preliminary studies were done towards understanding the mechanisms of MCH action on telost fish.

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"May 2003." Includes a list of publications by the author (journal articles related to thesis); and , copies of journal articles co-authored by the author. Includes bibliographical references (leaves 226-242) The solution structures of three peptides: Ala4Lys14-citopin 1.1 (amphipathic đ-helix); Gly15Gly19-caerin 1.1 (a less defined đ-helix); and, frenatin 3.1 (amphipathic đ-helix with a flexible c-terminal end) are presented in a discussion about structure/activity relationship

Thesis (Ph. D.)--Chemistry and Biochemistry, Georgia Institute of Technology, 2010. / Committee Chair: Bridgette Barry; Committee Member: David Collard; Committee Member: Ingeborg Schmidt-Krey; Committee Member: Jake Soper; Committee Member: Mira Josowicz. Part of the SMARTech Electronic Thesis and Dissertation Collection.