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Title: Chemical modification of nucleosides and chemical cross-linking of polynucleotides
Author: Hui, Cho Fat
ISNI:       0000 0001 3584 069X
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 1980
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The methylation of 5-hydroxycytidine with dimethyl sulphate or diazomethane in order to give 5-methoxycytidine was found to be unsuccessful. The methylation reactions gave two products, the 5-methoxycytidine and the dimethylated compound N4-methyl-5-methoxycytidine. The overall yields of these reactions were low and the two methylated products were difficult to be separated. When an attempt was made to synthesise N4-acetyl-5-hydroxycytidine, in order to protect the N-4 position from being methylated, it was found that N4-acetyl C was very labile towards the conditions employed in the hydroxylation reaction. Therefore, another method was employed for the synthesis of 5-methoxycytidine. The first stage of the synthesis of mo5C was the synthesis of 5-methoxyuracil, followed by the nucleoside synthesis. The next step after the nucleoside synthesis was the synthesis of 4-thio-5-methoxyuridine-2',3',5'-tri-0-benzoate, and finally, the conversion of the 4-thio group to the 4-amino group and the removal of the acyl protecting groups. The 5-methoxycytidine was phosphorylated to give 5-methoxycytidine-5'-diphosphate and polymerised with polynucleotide phosphorylase to give poly(mo5C). Poly(mo5C) formed a 1:1 complex with poly(I), and at neutral solution in the presence of 0.1 M sodium chloride, the Tm value was 64.5°. However, this double-stranded complex was ineffective as an interferon inducer. Attempts were made to synthesise alternating copoly(I-C) with polynucleotide phosphorylase, and were found to be unsuccessful. Alternating copoly(I-C) was synthesised with DNA-dependent RNA polymerase in the presence of template. Chemical cross-linking experiments of poly(I).poly(C) and alternating copoly(I-C), using a difunctional nitrogen mustard (methyl-1-N-bis(2-chloroathyl)amie hydrochloride) or 8-methoxypsoralen were carried out. In the case of poly(I).poly(C), no cross-linking was observed in both cross- linking reagents. Alternating copoly(I-C) was cross-linked by 8-methoxypsoralen, and the cross-linked and non-cross-linked species were separated by hydroxylapatite chromatography at room temperature. The effectiveness of this cross-linked polynucleotide as an interferon inducer was found to have decreased.
Supervisor: Not available Sponsor: British Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QD Chemistry