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Conformationally Co...
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Varghese, Oommen P.,1977-Uppsala universitet,Institutionen för biokemi och organisk kemi
(författare)
Conformationally Constrained Nucleosides : Design, Synthesis, and Biochemical Evaluation of Modified Antisense Oligonucleotides
Förlag, utgivningsår, omfång ...
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Uppsala :Acta Universitatis Upsaliensis,2007
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60 s.
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electronicrdacarrier
Nummerbeteckningar
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LIBRIS-ID:oai:DiVA.org:uu-8266
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ISBN:9789155469924
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https://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-8266URI
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Språk:engelska
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Sammanfattning på:engelska
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Klassifikation
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Ämneskategori:vet swepub-contenttype
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Ämneskategori:dok swepub-publicationtype
Serie
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Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Science and Technology,1651-6214 ;354
Anmärkningar
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This thesis is concerned with synthesis, structure and biochemical analysis of chemically modified oligonucleotides with potential therapeutic applications. The three types of chemical modifications described here are: (a) A North-East locked 1',2'-azetidine nucleoside (b) A North locked 2',4'-cyanomethylene bridged nucleoside and (c) A 2',4'-aza-ENA-T nucleoside. The synthesis of the 1',2'-azetidine fused nucleosides was described using two different approaches. A highly strained 2',4'-cyanomethylene locked nucleoside was synthesized but could not be converted to the phosphoramidite derivative due to instability during derivatization. The key cyclization step in the aza-ENA-T nucleoside synthesis gave rise to two separable diastereomers due to chirality at the exocyclic nitrogen. Conversion of diastereomer 55 to 56 occurred with a large free energy of activation (ΔG‡ = 23.4 kcal mol-1 at 298 K in pyridine-d5). Of the two isomers the equatorial NH product was more stable than the axial one due to reduced 1,3 diaxial interactions. As a result, all NH axial product was converted to the equatorial isomer during subsequent steps in the synthesis. NMR and ab initio experiments confirmed the North-East structure of the 1',2'-azetidine locked nucleoside and North conformation of aza-ENA-T locked nucleosides with a chair conformation of the piperidine ring.The amino modified nucleosides were incorporated into different positions of a 15mer oligonucleotide. The azetidine modified AONs did not form stable duplexes with complementary RNA (ΔTm ~-1 to -4 °C), but they performed better than previously synthesized isosequential 1',2'-oxetane modified oligonucleotides. The 2',4'-aza-ENA-T modified oligonucleotide, on the other hand, showed excellent target affinity with complementary RNA (ΔTm ~+4 °C). The azetidine and aza-ENA-T modified oligonucleotides showed significant stability in the presence of human serum and snake venom phosphodiesterase (3'-exonuclease) as compared to the unmodified native sequence. The singly modified 15mer oligonucleotides were also subjected to RNase H promoted digestion in order to evaluate their potential as effective antisense agents. The effective enzyme activity (kcat/Km) was found to be lower in the modified AONs due to reduced enzyme-substrate binding. However, the catalytic activity of RNase H with these modified-AON:RNA duplexes were higher than observed with the native duplex.
Ämnesord och genrebeteckningar
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NATURVETENSKAP Kemi hsv//swe
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NATURAL SCIENCES Chemical Sciences hsv//eng
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Organic chemistry
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chemically modified oligonucleotides
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azetidine
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aza-ENA-T
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cyanomethylene locked
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free energy of activation
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diaxial interactions
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chair conformation
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stable duplex
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human serum
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snake venom phosphodiesterase
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antisense agents
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RNase H
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Organisk kemi
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Chemistry
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Kemi
Biuppslag (personer, institutioner, konferenser, titlar ...)
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Engman, Lars
(preses)
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Stawinski, Jacek,ProfessorDepartment of Organic Chemistry, Stockholm University
(opponent)
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Uppsala universitetInstitutionen för biokemi och organisk kemi
(creator_code:org_t)
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