5 -deoxyribonucleotides

Cytosine was isolated from hydrolysis of calf thymus in 1894 and by 1903 its structure was known and it had been synthesized from 2-ethylthiopyrimidin-4(3H)-one. The acid hydrolysis of ribonucleic acid gives nucleotides, among which are two cytidylic acids, 2 -and 3 -phosphates of cytidine further hydrolysis gives cytidine itself, i.e. the 1-/3-D-ribofuranoside of cytosine, and thence cytosine. The deoxyribonucleic acids likewise yield deoxyribonucleotides, including cytosine deoxyribose-5 -phosphate, from which the phosphate may be removed to give cytosine deoxyriboside and thence cytosine. 

Methylcytosine (964 X = O) was synthesized in 1901 and its isolation from hydrolyzates of tubercule bacilli was reported in 1925. However, this was later shown to be incorrect and only about 1950 was it isolated by hydrolysis of the deoxyribonucleotide fractions from thymus, wheat germ and other sources (50MI21302). Nucleotides and a nucleoside of 5-methylcytosine are known. 

Et2AlCl, CH2CI2, 3 min, 70-85% yield.This method was used to remove the trityl group from various, protected deoxyribonucleotides. The TBDPS group is stable to these conditions. 

The most conspicuous use of iron in biological systems is in our blood, where the erythrocytes are filled with the oxygen-binding protein hemoglobin. The red color of blood is due to the iron atom bound to the heme group in hemoglobin. Similar heme-bound iron atoms are present in a number of proteins involved in electron-transfer reactions, notably cytochromes. A chemically more sophisticated use of iron is found in an enzyme, ribo nucleotide reductase, that catalyzes the conversion of ribonucleotides to deoxyribonucleotides, an important step in the synthesis of the building blocks of DNA. 

Nucleic acids are linear polymers of nucleotides linked 3 to 5 by phosphodi-ester bridges (Figure 11.17). They are formed as 5 -nucleoside monophosphates are successively added to the 3 -OH group of the preceding nucleotide, a process that gives the polymer a directional sense. Polymers of ribonucleotides are named ribonucleic acid, or RNA. Deoxyribonucleotide polymers are called deoxyribonucleic acid, or DNA. Because C-1 and C-4 in deoxyribonucleotides are involved in furanose ring formation and because there is no 2 -OH, only... 

The IDTr group was developed to protect the 5 -OH of deoxyribonucleotides and to increase the rate of intemucleotide bond formation through participation of... 

The benzoylformate ester can be prepared from the 3 -hydroxy group in a deoxyribonucleotide by reaction with benzoyl chloroformate (anhyd. Pyr, 20°,... 

Figure 28.1 Structures of the four deoxyribonucleotides and the four ribonucleotides.

Draw the complete structure of the deoxyribonucleotide sequence from which the mRNA codon in Problem 28.24 was transcribed. 

Deoxyribonucleic acid (DNA) (Section 28.1) The biopolymer consisting of deoxyribonucleotide units linked together through phosphate-sugar bonds. Found in the nucleus of cells, DNA contains an organism s genetic information. 

Polymer (Sections 7.10, 21.9, Chapter 31 introduction) A large molecule made up of repeating smaller units. For example, polyethylene is a synthetic polymer made from repeating ethylene units, and DNA is a biopolymer made of repeating deoxyribonucleotide units. 

Donahue, S.M., Brown, C.W., Scott, M.J., "Analysis of Deoxyribonucleotides with Principal Component and Partial Least-Squares Regression of UV Spectra after Fourier Processing", Appl. Spec. 1990 (44) 407-413. 

RNA is as suitable (if not more so) than DNA as a cleavage target [37]. In contrast to DNA, RNA is substantially less prone to oxidative cleavage [38] as a consequence of the higher stability of the glycosidic bond in ribonucleotides compared to that in deoxyribonucleotides. On the basis of the properties described in the introductory sections RNA is by contrast, much less stable to hydrolytic cleavage. For this reason the hydrolysis of the phosphate bond in this system can be successfully catalyzed not only by metal ions but also by ammonium ions. 

Figure 34-6. Regulation of the reduction of purine and pyrimidine ribonucleotides to their respective 2 -deoxyribonucleotides. Solid lines represent chemical flow. Broken lines show negative ( ) or positive ( ) feedback regulation.

Figure 36-16. The discontinuous poiymerization of deoxyribonucleotides on the lagging strand formation of Okazaki fragments during iagging strand DNA synthesis is illustrated. Okazaki fragments are 100-250 nt iong in eukaryotes, 1000-2000 bp in prokaryotes.

An alternative approach is to synthesize an artificial gene in the test-tube starting with the appropriate deoxyribonucleotides. This approach, which demands that the entire amino acid sequence be known, has been used to clone genes encoding proteins 200 amino acids long. 

Enzyme Taq polymerase (or some other enzyme) adds new deoxyribonucleotides during strand elongation. Taq is added to the assay at 1 unit per 50 qL of reaction mixture. 

The 3, 5 intemucleotide linkage is formed either by condensing die 3 -hydroxy group of an appropriately protected deoxyribonucleotide or -nucleoside with the 5 -phosphate of a deoxyribonucleotide (method a), or by condensing a 3 -phosphordiester with the 5 -hydroxy group of a nucleoside in a modified phosphortriester approach (method b). 

Using method a, oligodeoxyribonucleotides were synthesized from di- to deca-deoxyribonucleotides by means of mesitylenesulfonylimidazole and mesitylenesulfonyl-1,2,4-triazole. With triisoproylbenzenesulfonylimidazole die condensation took place more slowly.11121 Compared widi the corresponding arylsulfonyl chlorides, imidazolides induced intemucleotide condensation much more slowly, but caused no darkening of the reaction mixture, did not affect acid-sensitive bonds in trityl protected nucleotides, and did not sulfonate the 3 -hydroxy groups.11111 The reaction conditions were room temperature, 5—6 days, and pyridine as solvent.11111... 

Under certain circumstances DNA has both primer and template activities. For example, the addition of mononucleotides is to the 3 end of the growing DNA primer. This presents a problem with regard to how the other strand is synthesized. Biochemists have looked hard but unsuccessfully for an enzyme that can add deoxyribonucleotides onto the 5 end of DNA primers. Such a primer should contain a triphosphate on the hydroxyl group of the 5 end. Although a very active 5 -exonuclease, actually part of DNA polymerase I, has made the search for such an activated 5 end extremely difficult, investigators conclude that a polymerase able to use such a primer probably does not exist. On the contrary, good evidence suggests that the synthesis of both strands is by the known DNA poly-merases. 

The RNA oligonucleotides are complementary to a sequence on one of the strands of the DNA template and base pair with a portion of the DNA molecule. Subsequently, deoxyribonucleotides are covalently attached to the RNA primer. The synthesis of the primer itself is catalyzed by a special RNA polymerase called primase. Similar RNA polymerase-like enzymes are used to prime the synthesis of certain viral DNAs and eukaryotic DNA. 

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