Skip to main content
eScholarship
Open Access Publications from the University of California

Scholarly Works (5 results)

  • Thesis
  • Peer Reviewed
UC Irvine Electronic Theses and Dissertations (2022)

Chemical building blocks and models can serve as useful tools and surrogates for mimicking peptides in a biological system. In the following chapters of my thesis, I introduce two noncanonical amino acid building blocks that I have synthesized for incorporation into peptides and evaluate their secondary structure formation or biological activity (chapter 1 and chapter 3). I will also describe the use of chemical models derived from the β-amyloid peptide Aβ to investigate and probe the chiral selectivity in β-sheet assembly in aqueous solution (chapter 2). These three chapters of my thesis represent my passion for organic synthesis and NMR spectroscopy, and are linked by a theme of using building blocks and models to investigate peptide folding, assembly, and activity.

Chapter 1 describes the design, synthesis, and evaluation of an improved turn structure for inducing β-hairpin formation in peptides. This project builds upon a previous finding from our laboratory that δ-linked-ornithine (δOrn) can be used as suitable turn unit for stabilizing macrocyclic β-sheet peptides. Development of β-turn units as peptidomimetic tools are important for stabilizing the secondary structure of peptides and have led to the design of biomaterials and potential biomedical therapeutics. Chapter 1 details the development and characterization of γ-methylornithine — a methylated version of δOrn — and shows its propensity to induce a β-hairpin conformation in comparison to the established turn, D-Pro-Gly. The utility provided by this improved turn structure will add to the existing toolbox of ways to create stabilized macrocyclic β-sheet peptides.

Chapter 2 seeks to answer a basic question: do enantiomeric β-sheet peptides prefer to self-assemble to form pleated β-sheets or co-assemble to form rippled β-sheets? In the past decade, a theory has emerged that enantiomeric β-sheets prefer to co-assemble in a heterochiral fashion to form rippled β-sheets. Our laboratory has previously found that enantiomeric β-sheet pentapeptides strongly prefer to self-assemble. To address this discrepancy, equimolar amounts of L- and D-chemical model peptides derived from Aβ were mixed and studied by solution-phase NMR spectroscopy. These NMR spectroscopic studies reveal that β-sheet peptide models derived from Aβ prefer to self-assemble in a homochiral fashion in aqueous solution. Chapter 2 ends with concluding remarks about our interpretation of the different forces that drive the formation of heterochiral rippled β-sheets vs. homochiral pleated β-sheets.

Chapter 3 is an ongoing collaborative project with members of the lab, on the synthesis and stereochemical determination of Novo29, a new Gram-positive peptide antibiotic discovered by Novobiotic Pharmaceuticals. Novo29 is an 8-residue peptide that contains a noncanonical hydroxyasparagine amino acid at position 5; the stereochemistry of this amino acid is undetermined. Chapter 3 begins with the syntheses of two diastereomers of hydroxyasparagine containing either the R,R or the R,S stereochemistry at the α- and β-positions. Incorporation of the hydroxyasparagine amino acids into peptides enabled the stereochemical determination of the natural Novo29 through NMR spectroscopic studies and functional correlation assays. At the end of chapter 3, a 1.13 Å X-ray crystallographic structure of the synthetic peptide containing the R,S stereochemistry is presented and discussed in further detail. Future directions for this project will aim to generate a molecular model of the peptide containing R,R stereochemistry in order to provide an explanation into the potential mechanism of action of the natural Novo29 antibiotic.

    Cover page: Chemical Building Blocks and Models for Studying Peptides
    • Article
    • Peer Reviewed
    UC Irvine Previously Published Works (2021)
    Although β-hairpins are widespread in proteins, there is no tool to coax any small peptide to adopt a β-hairpin conformation, regardless of sequence. Here, we report that δ-linked γ(R)-methyl-ornithine (δ MeOrn) provides an improved β-turn template for inducing a β-hairpin conformation in peptides. We developed a synthesis of protected δ MeOrn as a building block suitable for use in Fmoc-based solid-phase peptide synthesis. The synthesis begins with l-leucine and affords gram quantities of the Nα -Boc-Nδ -Fmoc-γ(R)-methyl-ornithine building block. X-ray crystallography confirms that the δ MeOrn turn unit adopts a folded structure in a macrocyclic β-hairpin peptide. CD and NMR spectroscopy allow comparison of the δ MeOrn turn template to the δ-linked ornithine (δ Orn) turn template that we previously introduced and to the popular d-Pro-Gly turn template. These studies show that the folding of the δ MeOrn turn template is substantially better than that of δ Orn and is comparable to d-Pro-Gly.
      Cover page: An Improved Turn Structure for Inducing β-Hairpin Formation in Peptides.
      • Article
      • Peer Reviewed
      UC Irvine Previously Published Works (2022)
      In 1953, Pauling and Corey postulated "rippled" β-sheets, composed of a mixture of d- and l-peptide strands, as a hypothetical alternative to the now well-established structures of "pleated" β-sheets, which they proposed as a component of all-l-proteins. Growing interest in rippled β-sheets over the past decade has led to the development of mixtures of d- and l-peptides for biomedical applications, and a theory has emerged that mixtures of enantiomeric β-sheet peptides prefer to co-assemble in a heterochiral fashion to form rippled β-sheets. Intrigued by conflicting reports that enantiomeric β-sheet peptides prefer to self-assemble in a homochiral fashion to form pleated β-sheets, we set out address this controversy using two β-sheet peptides derived from Aβ17-23 and Aβ30-36, peptides 1a and 1b. Each of these peptides self-assembles to form tetramers comprising sandwiches of β-sheet dimers in aqueous solution. Through solution-phase NMR spectroscopy, we characterize the different species formed when peptides 1a and 1b are mixed with their respective d-enantiomers, peptides ent-1a and ent-1b. 1H NMR, DOSY, and 1H,15N-HSQC experiments reveal that mixing peptides 1a and ent-1a results in the predominant formation of homochiral tetramers, with a smaller fraction of a new heterochiral tetramer, and mixing peptides 1b and ent-1b does not result in any detectable heterochiral assembly. 15N-edited NOESY reveals that the heterochiral tetramer formed by peptides 1a and ent-1a is composed of two homochiral dimers. Collectively, these NMR studies of Aβ-derived peptides provide compelling evidence that enantiomeric β-sheet peptides prefer to self-assemble in a homochiral fashion in aqueous solution.
        Cover page: Enantiomeric β-sheet peptides from Aβ form homochiral pleated β-sheets rather than heterochiral rippled β-sheetsCreative Commons 'BY' version 4.0 license
        • Article
        • Peer Reviewed
        UC Irvine Previously Published Works (2020)
        This paper describes the chemical synthesis, X-ray crystallographic structure, and antibiotic activity assay of lactam analogues of teixobactin and explores ring-expanded analogues of teixobactin with β3-homo amino acids. Lactam analogues of teixobactin containing all four stereoisomers of aza-threonine at position 8 were synthesized on a solid support from commercially available stereoisomeric threonine derivatives. The threonine stereoisomers are converted to the diastereomeric aza-threonines by mesylation, azide displacement, and reduction during the synthesis. d-Aza-Thr8,Arg10-teixobactin exhibits 2-8-fold greater antibiotic activity than the corresponding macrolactone Arg10-teixobactin. Azateixobactin analogues containing other stereoisomers of aza-threonine are inactive. A dramatic 16-128-fold increase in the activity of teixobactin and teixobactin analogues is observed with the inclusion of 0.002% of the mild detergent polysorbate 80 in the MIC assay. The X-ray crystallographic structure of N-Me-d-Gln4,d-aza-Thr8,Arg10-teixobactin reveals an amphipathic hydrogen-bonded antiparallel β-sheet dimer that binds chloride anions. In the binding site, the macrolactam amide NH groups of residues 8, 10, and 11, as well as the extra amide NH group of the lactam ring, hydrogen bond to the chloride anion. The teixobactin pharmacophore tolerates ring expansion of the 13-membered ring to 14-,15-, and 16-membered rings containing β3-homo amino acids with retention of partial or full antibiotic activity.
          Cover page: Design, Synthesis, and Study of Lactam and Ring-Expanded Analogues of Teixobactin.
          • Article
          • Peer Reviewed
          UC Irvine Previously Published Works (2023)
          This paper describes the synthesis and stereochemical determination of Novo29 (clovibactin), a new peptide antibiotic that is related to teixobactin and is active against Gram-positive bacteria. Novo29 is an eight-residue depsipeptide that contains the noncanonical amino acid hydroxyasparagine of hitherto undetermined stereochemistry in a macrolactone ring. The amino acid building blocks Fmoc-(2R,3R)-hydroxyasparagine-OH and Fmoc-(2R,3S)-hydroxyasparagine-OH were synthesized from (R,R)- and (S,S)-diethyl tartrate. Novo29 and epi-Novo29 were then prepared by solid-phase peptide synthesis using these building blocks. Correlation with an authentic sample of Novo29 through 1H NMR spectroscopy, LC-MS, and in vitro antibiotic activity established that Novo29 contains (2R,3R)-hydroxyasparagine. X-ray crystallography reveals that epi-Novo29 adopts an amphiphilic conformation, with a hydrophobic surface and a hydrophilic surface. Four sets of epi-Novo29 molecules pack in the crystal lattice to form a hydrophobic core. The macrolactone ring adopts a conformation in which the main-chain amide NH groups converge to create a cavity, which binds ordered water and acetate anion. The amphiphilic conformation of epi-Novo29 is reminiscent of the amphiphilic conformation adopted by the related antibiotic teixobactin and its derivatives, which contains a hydrophobic surface that interacts with the lipids of the bacterial cell membrane and a hydrophilic surface that interacts with the aqueous environment. Molecular modeling suggests that Novo29 can adopt an amphiphilic conformation similar to teixobactin, suggesting that Novo29 may interact with bacteria in a similar fashion to teixobactin.
            Cover page: Synthesis and Stereochemical Determination of the Peptide Antibiotic Novo29.Creative Commons 'BY' version 4.0 license
            Top