Design and synthesis of unnatural fluorescent triazolyl amino acids and constrained molecular scaffolds and their application in peptidomimetics" was carried out by me under the supervision of Dr. Thesis Submitted (June)] Title: “Design and Synthesis of Unnatural Fluorescent Aminoacidol and Triazole Restrained Molecular Scaffold and Their Application in Peptidomimetics.
CHAPTER 1: APPLICATIONS OF UNNATURAL AMINO ACIDS AND
Conformational constrained amino acids/scaffolds limit the rotation of amide bond, side chain modified amino acids can limit conformational flexibility via steric interactions and help to adopt a folded conformation without drastically changing the stereo-electronic properties. To this end, several unnatural amino acids (UNAA)/amino acid scaffolds with novel functionalities have been designed and incorporated within a protein's framework with the aim of expanding the genetic code or having enzymatically stable peptide-based drug candidates.
CHAPTER 2: STUDIES ON THE SYNTHESIS AND PHOTOPHYSICAL PROPERTIES OF FLUORESCENT UNNATURAL TRIAZOLYL AMINO
The synthesized fluorescent amino acids exhibit interesting solvatochromic features and/or intramolecular charge transfer (ICT), as evidenced by the UV-visible, fluorescent photophysical properties and DFT/TDDFT calculation. The HOMO-LUMO distribution shows that the emission states of some amino acids are characterized by more significant electron redistribution between the triazolyl group and the associated aromatic chromophores, leading to modulated emission properties.
CHAPTER 3: SYNTHESIS, CONFORMATION AND STUDY OF PHOTOPHYSICS OF TRIAZOLYL DONOR–ACCEPTOR UNNATURAL
Thus, we were interested in β-turn peptidomimetics capable of displaying FRET interaction between two terminally positioned fluorescent unnatural amino acids. Förster resonance energy transfer (FRET) was also established in the conceptually designed new unnatural β-turn peptide containing a new class of fluorescent unnatural donor/acceptor amino acids.
We observed that a designed triazolo-amino acid with constrained backbone angles (i) and (i+1) would be expected to induce folded conformations in linear peptides. Exploration of the tourmimetics and the sequence-specific DNA binding event for tetra-amides of this molecular scaffold may lead to the creation of a new family of distamycin analogs. The readily available aliphatic triazoloamino acid.
CHAPTER 5: DESIGN AND SYNTHESIS OF AROMATIC TRIAZOLO AMINO ACID AS CONFORMATIONALLY CONSTRAINED MOLECULER
An examination of the photophysical properties of the trichromophoric fluorescent pentapeptide established the basic concept of double-door entry into excimer emission. This study will provide fundamental guidelines to design such a conceptual fluorescent probe of the dual-gate entry system in excimer emission.
CHAPTER 6: APPLICATION OF AROMATIC TRIAZOLYL AMINO ACID SCAFFOLD AND ITS MONO- AND BIS-PYRENYL AMIDES AS
Therefore, we took on the task of detecting ethanol using the aromatic triazolo-amino acid scaffold ArTAA. Monopyrenylamide aromatic triazoloamino acid Bis-pyrenyl-bis-amide aromatic triazoloamino acid Per.
APPLICATIONS OF UNNATURAL AMINO ACIDS
CHAPTER 2: STUDIES ON THE SYNTHESIS AND PHOTOPHYSICAL PROPERTIES OF FLUORESCENT
Studies On the Interaction of Amino Acid TPer Ala Do (2.78) with BSA
CHAPTER 3: SYNTHESIS, CONFORMATION AND STUDY OF PHOTOPHYSICS OF TRIAZOLYL DONOR–ACCEPTOR
Conformational Study of Peptides 4.102 and 4.103 Using CD, IR, NMR, Spectroscopic Techniques and Macromodel Calculations. Conformational Study of Peptides 5.42 and 5.43 Using CD, IR, NMR, Spectroscopic Techniques and Macromodel Calculations.
CHAPTER 6: APPLICATION OF AROMATIC TRIAZOLYL AMINO ACID SCAFFOLD AND ITS MONO- AND BIS-
Introduction
However, in all organisms, the building blocks of all translated proteins are the same 20 natural amino acids. The following sections of this chapter will represent a critical survey of the applications of conformationally restricted unnatural amino acids and small molecule/non-peptide isosteres in the context of peptidomimetics and the generation of functional peptides with novel photophysical properties.
Need for Unnatural Amino Acids (UNAAs)
Unnatural amino acids (uNAAs) can also be used as a probe for the structural and functional study of proteins by biophysical techniques such as nuclear magnetic resonance (NMR), electron paramagnetic resonance (EPR) and infrared (IR) spectroscopy.24j . All three of these major research efforts toward the development of unnatural amino acids are briefly outlined below.
Unnatural Amino Acids for Genetic Incorporation
Summary of the key reagent required for the incorporation of unnatural amino acids into proteins using nonsense codon suppression. Five-base codon-anticodon pairs for introducing unnatural amino acids into proteins.
Fluorescent Unnatural Amino Acids for Protein Monitoring
- Fluorescent Unnatural Amino Acids: Tryptophan/Tyrosine Mimics
- Fluorescent Unnatural Amino Acids Carrying Polyaromatic Side Chain Unnatural amino acids with polyaromatic side chain are useful fluorescent probe
- Coumaryl Fluorescent Nnnatural Amino Acids
- Fluorescent Unnatural Amino Acids: Microenvironment Sensitivity
- Fluorescent Unnatural Amino Acid: Sensing Metal Ion
Sisido et al., efficiently incorporated several non-natural amino acids with aromatic side chain (Figure 1.16). Another polarity-sensitive fluorescent unnatural amino acids L-Anap was reported and genetically encoded by Wang et.
Unnatural Amino Acids as Small Molecule Scaffold for Peptidomimetics
- Proline Analogue as β-Turns Inducer
- Sugar Amino Acids (SAAs) as a Turn Inducer: Leu-Enkephalin Analogue The five membered sugar ring carrying one carboxyl and one amino functional
- Cyclopentane-Based γ‑ Amino Acid as a Molecular Backbone
- Pyrrole and Furan Based Peptidomimetics
- Enediynyl Amino Acid as β- Turn Nucleator
- Norbornene Containing Peptide: β-Sheet Nucleator
- Dibenzofuran-Based Molecular Scaffold
- Biphenyl Scaffold as a Peptide Backbone
- Pyridine-Based Molecular Scaffolds
- Epindolidione as a β-Sheet Nucleator
- Triazole Scaffold as peptidomimetics
The peptide 1.200 also exhibited parallel β-sheet structure in DMSO-d6.96. The adopted conformations of the peptide where norbornene was placed in the peptide backbone clearly suggested that this scaffold acts as a good β-sheet nucleator. The formation of such head-to-tail dimeric structures is very unique and similar to the β-strand structure found in many peptides (Figure 1.42).
Summary and Future Prospect
Sze group et al. reported a new class of 1,4-triazole-based peptidomimetics that self-dimerize from head to tail and form organogel in both nonpolar and protic organic solvent. On the other hand, the generation of foldamer structures bearing isosteres of amino acid side chains and exhibiting a secondary framework such as a β-hairpin/β-sheet or α-helix conformation holds promise for the next generation of therapeutics in medicine.
Introduction
Methods of 1,2,3-triazole synthesis
Schematic representation of a fluorogenic Cu(I) catalysed click reaction
Scheme 2.3. Fluorogenic CuAAC reaction to produce highly fluorescent coumarin derivatives starting from weakly fluorescent coumarine
Click Chemistry and Drug Design
Therefore, click chemistry in this pathway has gained attention in synthesizing a combinatorial library of compounds that can be screened after studying the interactions with the unexplored targets. Primarily, click chemistry-based drug discovery falls into three categories (1) high throughput screening, (2) fragment-based drug discovery, and (3) dynamic template-assisted strategies in fragment-based drug discovery.
Synthesis and structure of the triazole modified unnatural amino acids
- Oligo-Triazole Based Peptide Nucleic Acid Analogue (T-PNA)
- Some Examples of Unnatural and Fluorescent Amino Acids
- Background
- Objective
36 Interest in the design and application of new unnatural amino acids is increasing universally. Furthermore, we imagined that the fluorescent amino acids could also be useful in the study of.
Result and Discussion
- Synthesis of Triazolyl Donor/Acceptor Amino Acids
Synthetic scheme for donor-acceptor triazolyl unnatural amino acids
Spectral Characterization of Few Representative Triazolyl Unnatural Fluorescent Amino Acids
As a representative example, the structural assignments of two protected triazolyl amino acids 2,074 and 2,079 are shown in Figure 2.9. For amino acid 2,074 (TPhenAlaDo), the triazolyl hydrogen appeared as a singlet at the characteristic position of 7.94.
Study of photophysical properties
UV-visible absorption and fluorescence emission spectra of amino acids in various organic solvents; (a-b) for 2.72 (TDMBAlaDo) and (c-d) for 2.73 (TMNapAlaDo). UV-visible absorption and fluorescence emission spectra of amino acids in various organic solvents; (a-b) for 2.76 (TPyAlaDo) and (c-d) for 2.77 (TBAPyAlaDo).
Study of Possible Photophysical Interaction Among a Donor/Acceptor Pair of Triazolyl Unnatural Fluorescent Amino
This observation clearly indicated a quenching of the fluorescence of TCNBAlaAc in the presence of TPhenAlaDo via a dynamic quenching process, as the static mode of quenching was excluded from the UV–visible spectroscopic study. As the concentration of the FRET acceptor (TPhenAlaDo) increases, the fluorescence of the FRET donor (TCNBAlaAc) decreases (Figure 2.18c).
Theoretical Calculations
The emission states of all the donor triazole amino acids were found to be locally excited (LE) states, except for the case of amino acid 2.70 (TNDMBAlaDo) as suggested by the HOMO−LUMO distributions. In the case of amino acid 2.70, the emission state was characterized with more significant electron redistribution, i.e., ICT feature.
Study of Interaction of Amino Acid 2.78 ( TPer Ala Do ) with BSA Protien
The amino acid sequence of the BSA protein was observed via the NCBI website, http://www.ncbi.nlm.nih.gov/protein/CAA76847.1. The 3D model of the BSA protein was built using the 3D structure 1AO6 chain 'A' as a template using the ESyPred3D11 web server. The binding of the TPer moiety in the hydrophobic pocket of BSA was also supported by a molecular docking calculation with the Autodoc program59, which clearly showed that the TPer moiety of the probe 2.78 remained surrounded by hydrophobic amino acids of the hydrophobic pocket of subdomain IB of the site. I of BSA (Figure 2.23).
Conclusion
Experimental Section
- General Experimental
- Synthesis and Characterization
- General procedure for the synthesis of triazolyl donor/acceptor amino acids by [3+2] cycloaddition reaction
- Photophysical Studies of the Amino Acids
- Studies On the Interaction of Amino Acid TPer Ala Do (2.78) with BSA
The reaction mixture was stirred until the total starting material was consumed, monitored by TLC, then poured into water and CH 2 Cl 2 (20 mL each). The organic layer was washed with water twice, dried under Na 2 SO 4 and then evaporated. The absorbance values of the sample solutions were measured in the wavelength regime of 200-550 nm.
- References
- Introduction: Peptidomimetics and Their Importance
- The Approaches to β-Turn Peptidomimetic
- β-Turn Peptidomimetics
- Fluorescently Labelled Peptides/Proteins and Application of FRET Process
Most of the β-turn structures are formed by creating intramolecular hydrogen bonding Torsion angles for classical β-turns. Schematic representation of type II β-turn peptides with unnatural amino acid, Aib (α-amino isobutyric acid) in the middle of the peptide strand.
T inact
Background
From a study of the literature, it appears that there are a very limited number of fluorescent amino acids that have been synthesized or genetically encoded or incorporated into a short peptide to create fluorescent peptides that could adopt/mimicking a particular protein secondary structure and can exhibit very interesting photophysical properties. Additionally, a limited number of fluorescent amino acids were incorporated into short peptides to study protein-protein interaction.
Objective
A study of the possible photophysical interaction between the donor/acceptor triazolyl unnatural amino fluorescent acid pair, as already described in Chapter 2, Section 2.8, showed that the UV-visible titration of a solution of the acceptor triazolyl amino acid TCNBAlaAc (2.79) with increasing concentration of the donor triazolyl amino acids, TPhenAlaDo (2.74) showed characteristic absorptions of the donor and acceptor amino acids, indicating that there was no possibility of complexation by charge transfer in the ground state (Figure 2.18). Exploring the photophysics of other possible fluorescent donor/acceptor amino acid pairs is a future scope of this dissertation.
Results and Discussion
- Synthesis of Triazolyl Donor/Acceptor Amino Acid Decorated Tripeptides To synthesize our designed tripeptides decorated with our synthesized
Furthermore, it was observed that emission spectra of TCNBAlaAc overlapped significantly with the absorption spectra of TPhenAlaDo, indicating a possibility of quenching of emission of TCNBAlaAc via a dynamic quenching mode, FRET. Therefore, this donor/acceptor amino acid can potentially act as a FRET pair if they can be incorporated into a peptide construct, where the acceptor triazolyl amino acid TCNBAlaAc would act as a FRET donor and the donor amino acid TPhenAlaDo would act as a FRET acceptor.
Synthetic procedure of triazolyl decorated donor/acceptor tripeptides
- Spectral Characterisation of the Synthesized Tripeptides
- H (Ala)
- Study of Photophysical Properties of Synthesized Tripeptides
- Study of Förster Resonance Energy Transfer (FRET) in Peptide 3.37
- Solvent Assisted Modulation of FRET Emission
- Conclusion
- Experimental Section
Then the reaction mixture was poured into water and CH 2 Cl 2 (20 mL each). The organic layer was washed with water twice, dried over Na 2 SO 4 and then evaporated. The organic layer was washed with brine, dried over Na 2 SO 4 . The title trizolyl derivative of dipeptides 3.47 and 3.48 was separated by column chromatography and characterized.
In the chemistry and biochemistry of amino acids, peptides and proteins; Weinstein, B., ed.; Marcel Dekker: New York, 1983, VII, 267.
Introduction
Peptidomimetics: Design and Classifications
It is the structural mimetic of a natural substrate with all the functionalities in a well-defined spatial orientation. These are both structural and functional imitations, which can be used via the introduction of a scaffold with a different structure to that of the substrate.
Approaches to Peptidomimetic Design
- Introduction of Global Restrictions
- Molecular Scaffolds Mimicking the Peptidic Backbone: Peptidomimetic Scaffolds
- Molecular Scaffolds Mimicking/Inducing The β-Turns
Bio isosteres of the peptide bond. i) Side chain isosteres: In addition, side chain modifications have also been introduced to investigate pharmacophoric steric and electronic interactions. The purpose of introducing local modifications around side chains is mainly to modulate the conformational profile of the peptide.
The Triazole Ring as a Peptidomimetic Scaffold
- Triazole-Containing Peptidomimetics
- Triazoles as β-Turn Inducers (a Small Molecule Scaffold/Dipeptide Isosteres/Dipeptide Mimic) in Secondary Structures
Comparison between a β-turn and β-turn peptidomimetics containing the triazole ring connecting the two peptide β-strands. Pan and his colleagues64 therefore used the triazole ring as an amino acid substitution to obtain a cyclic RGD tripeptide, cyclo[-Arg-Gly-Asp-(triazole)-Gly-Xaa-] via 'click chemistry' (Figure 4.13b) .
Background
Objective
-Leu-TPhenAlaDo-CO2Me), in which two fluorescent unnatural triazolo-amino acids, (TPyAlaDo and TPhenAlaDo), are at the two arms of the scaffold via an intervening leucine in each arm. d) Study of fluorescent photophysics of the pentapeptide and photophysical interaction, if any, between the terminal fluorescent amino acids.
Result and Discussion
- Synthesis of Aliphatic Triazolyl Amino Acid Scaffold (4.100, Al TAA) and Corresponding Peptides
Synthesis of aliphatic triazoly amino acid scaffold (4.100, Al TAA)
