Please use this identifier to cite or link to this item: http://localhost:8081/xmlui/handle/123456789/537
Authors: Kukreti, Shrikant
Issue Date: 1989
Abstract: Model systems involving nucleic acid and protein consti tuents have been widely used to obtain information on the specific interaction involved in protein-nucleic acid associa tions. The advantage to carry out this type of studies is the possibility to investigate interactions of specific nucleotide sequence of DNA with peptide chain with controlled amino acid composition, sequence, chain lengths and predictable conformation, which results in considerable simplification of the system as compared to the natural protein-DNA systems. Stacking of aromatic amino acids with nucleic acid bases is found to exist in the crystal structure of nucleoside peptide 5-[N-(L-phenylalanyl)-amino] uridine. Tyrosine, phenyla lanine and tryptophan are found to stack in complexes of prokaryotic ribonuclease T with 2'-GMP, gene 5 protein of phase fd with (dA)5, lac repressor with operator etc. The present study has been undertaken to investigate stacking interactions using model systems with a view to find base/base sequence specificity and conformational changes induced on stacking. We have studied interaction of Tyr, Trp, Phe containing tri- and tetrapeptide with d-CpG d-GpC, d-GpCpGpC and d-CpCpGpG. One-dimensional NMR is used to find chemical shifts due to stacking, changes in Tm etc. while two-dimen sional NMR techniques-J-correlated spectroscopy (COSY) and nuclear overhauser enhancement spectroscopy (NOESY) serve as methods to assign all proton NMR signals unambiguously and determine solution conformation such as sugar pucker, helix sense, glycosidic bond rotation and interproton dis tances. We also present results of theoretical energy calcula tions made on stacking of Tyr, Trp, Phe with C, G bases, A-T and C-G base pairs and intercalation between C-G and G-C base pairs. Classical potential function has been used to estimate conformational energy. Total interaction energies are calculated as sum of electrostatic, dispersion, polari zation and repulsion terms. The upfield shifts of aromatic ring proton resonances of Trp, Tyr and Phe indicate stacking. The interaction is more prominent with double-helix than with single-stranded DNA. Practically in all cases DNA is double-helix with and without binding, sugar conformations are mostly Ol'-endo, glycosidic bond rotations are anti/high anti. Interproton NOE's show most of the peptides are either in close proximity or stack with nucleotides. Among oligopeptides, the binding of d-GpCpGpC with Lys-Tyr-Lys is strongest. The binding of d-CpG with various aromatic amino acids of peptide have been found to be in the order Trp > Tyr > Phe. From theoretical studies, the interaction energies for model system d-CG with aromatic amino acids are in the same order and thus are in agreement with NMR results that we have obtained. From our studies, we have thus shown that there is a specified type of base/base sequences specificity in nucleic acid-protein interactions. This may be the reason for specific recognition of binding site of nucleic acid by proteins.
Other Identifiers: Ph.D
Research Supervisor/ Guide: Barthwal, Ritu
metadata.dc.type: Doctoral Thesis
Appears in Collections:DOCTORAL THESES (Bio.)

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