Certain aspects of polynucleotide biosynthesis

Part I. 1. The ability of extracts of Ehrlich ascites carcinoma to phosphoralate deoxyribonucleoside monophosphates was studied with a view to using this system as a means of preparing the deoxyribonucleoside triphosphates on a large scale. 2. The extracts were found to be able to convert dAMP to dATP in 60% - 80% yield, dGMP to dGTP in 50% - 60% yield, dCMP to dCTP in about 15% yield TMP to TTP in about 1% yield. 3. A method for the preparation of dATP and dGTP on an large seals, using extracts of Ehrlich ascites carcincma, is described. Part II. 1. TTP was prepared from TMP in 18% yield by the method described by Smith and Khorana for the preparation of ATP from AMP. 2. An investigation of the purification led to several improvements. 3. This improved method (Method I) was used in the isolation of TTP-H3 prepared from TMP-H3 by the method of Smith and Khorann. An overall yield of 36% was obtained. 4. The effect on the yield of deoxyribonucleoside triphosphate of deoreasing the ratio of the deoxyribonucleo-side memophosphate to the other constituents of the reaction mixture, was investigated. By using reaction mixture, involving twice and three times the amounts of the reactants used by Smith and Khorana for the same amount of deoxyribonucleoside monophosphate, greatly improved yields of the deoxyribonucleoside triphosphates were obtained. 5. The effect on the yield of deoxyribonucleoside triphosphate of reacting the deoxyribonuclcoside monophosphate in phosphate of reacting the deoxyribonuclcoside monophasphate in the from of the tri-n-butylammonium salt instead of an the free acid was also studied. Still futher improvements in the yields of the deoxyribonucleoside triphosphates were obtained. TMP was converted to TTP in 92% yield, dAMP to dATP in 93% yield, dGMP to dGTP in 92% yield and dCMP to dCTP in 73% yield. 6. A charcoal column procedure for the simultaneous removal of lithium chloride and inorphanic polyphosphates from the deoxyribonucleoside triphosphate solution obtained after anion exchange chromatography was developed and incorporated in Method I giving Method II. 7. TMP32 was prepared from thymidine by the methods of Rurwits and Tener. 8. The TMP32 thus obtained was converted, via the trin-butylammnium salt, to TTP in 90% yield and the TTP thus formed isolated by Method II in 70% yield. The specific activity of the TTP thus prepared was 1.0 x 1.5 x 10