Nucleic acids phosphodiester groups

Another important group of hydrolytic enzymes are phospho- and cyclophosphodiesterases. They catalyze the hydrolysis of phospho-diester bonds and many of the most relevant biological substrates are nucleic acids. Phospholipase C and D are also important examples. Initial attempts to measure phosphodiesterase activity placed a phosphodiester between a fluorophore and a quencher and the probe was tested in vitro [146], This system was slightly modified by Caturla and used for the identification of catalysts with phosphodiesterase activity [147], More recently, Nagano and co-workers used a coumarin donor and fluorescein as a FRET... 

In RNA, the base T found in DNA is replaced by uracil, which is similar in structure to T, but lacks the methyl group. The nucleotides in nucleic acids are linked by phosphodiester bonds between the 3 -hydroxyl of one nucleoside and the 5 -hydroxyl of the sugar of its neighbour in the sequence, as was first shown by Alexander Todd3 in 1952 (Figure 4.13). 

Nucleic acids are polymers of nucleotides joined by 3, 5 -phosphodiester bonds that is, a phosphate group links the 3 carbon of a sugar to the 5 carbon of the next sugar in the chain. Each strand has a distinct 5 end and 3 end, and thus has polarity. A phosphate group is often found at the 5 end, and a hydroxyl group is often found at the 3 end. 

A nucleotide consists of a heterocyclic base linked to a sugar (ribose or deoxyribose) and a phosphate group also linked to the sugar (Figure 10.6). Nucleic acids are polymers of nucleotides linked together by phosphodiester bonds (Figure 10.7). The enzymes that catalyse the breakdown of nucleic acids to nucleotides are nucleases. 

The nucleic acid polymer is formed when the nucleotides attach to one another through phosphodiester bonds, which connect the 3 -OH group of one nucleotide to the 5 -OH group of another nucleotide through the phosphate group. The order of the nucleotides in the chain is the primary structure of the DNA or RNA molecule, and it can be represented in short-hand notation with only the base pair designation... 

FIGURE 8-7 Phosphodiester linkages in the covalent backbone of DNA and RNA. The phosphodiester bonds (one of which is shaded in the DNA) link successive nucleotide units. The backbone of alternating pentose and phosphate groups in both types of nucleic acid is highly polar. The 5 end of the macromolecule lacks a nucleotide at the 5 position, and the 3 end lacks a nucleotide at the 3 position. 

All the phosphodiester linkages have the same orientation along the chain (Fig. 8-7), giving each linear nucleic acid strand a specific polarity and distinct 5 and 3 ends. By definition, the 5 end lacks a nucleotide at the 5 position and the 3 end lacks a nucleotide at the 3 position. Other groups (most often one or more phosphates) may be present on one or both ends. 

Nucleotides are the building blocks of nucleic acids their structures and biochemistry were discussed in chapter 23. When a 5 -phosphomononucleotide is joined by a phosphodiester bond to the 3 -OH group of another mononucleotide, a dinucleotide is formed. The 3 -5 -linked phosphodiester intemucleotide structure of nucleic acids was firmly established by Lord Alexander Todd in 1951. Repetition of this linkage leads to the formation of polydeoxyribonucleotides in DNA or polyribonucleotides in RNA. The structure of a short polydeoxyribonucleotide is shown in figure 25.3. The polymeric structure consists of a sugar phosphate diester backbone with bases attached as distinctive side chains to the sugars. 

Polynucleotide polymerases, or nucleotidyl transferases, are enzymes that catalyze the template-instructed polymerization of deoxyribo- or ribonu-cleoside triphosphates into polymeric nucleic acid - DNA or RNA. Depending on their substrate specificity, polymerases are classed as RNA- or DNA-dependent polymerases which copy their templates into RNA or DNA (all combinations of substrates are possible). Polymerization, or nucleotidyl transfer, involves formation of a phosphodiester bond that results from nucleophilic attack of the 3 -OH of primer-template on the a-phosphate group of the incoming nucleoside triphosphate. Although substantial diversity of sequence and function is observed for natural polymerases, there is evidence that many employ the same mechanism for DNA or RNA synthesis. On the basis of the crystal structures of polymerase replication complexes, a two-metal-ion mechanism of nucleotide addition was proposed [1] during this two divalent metal ions stabilize the structure and charge of the expected pentacovalent transition state (Figure B.16.1). 

In a given phosphodiester bond, hydrolytic enzymatic cleavage can occur at two locations, indicated by p and d in Figure 10.16. The former is proximal with respect to the 3 -OH group the latter is distal with respect to the 3 -OH. Enzymes that catalyze the hydrolysis of nucleic acids are nucleases (see Table 10.2). Exonucleases remove nucleotides (or nucleosides) from either the 5 or the 3 end of the polynucleotide. These are specific for either the p or the d bond. Thus, an exo-... 

Recently, several nucleophilic reagents have been used to establish the mode of action of the metabolites of polycyclic aromatic hydrocarbons (PAH). Among them, several phosphodiesters have been examined to clarify the possibility of reaction of PAH epoxides with the phosphate groups(P-alkylation) of nucleic acids (22). In this context we have studied the reaction of 3,4-epoxyprecocene II with dibenzyl phosphate under a variety of conditions. In all cases, instead of the formation of phenol or phosphotriesters observed with PAH epoxides, we obtained predominantly dimer XI. This compound was also the main component of the mixtures obtained by reaction of the above precocene epoxide with other acid catalysts, along with dimers XII and XII. Dimer XII was formed almost exclusively by thermal treatment. The structure and configuration for compound XII has been established by spectral and X-ray diffraction analyses (23). 

We described an unusual oligomer whereby a-amino acids were converted into a-hydroxy acids by oxidative deamination with retention of stereochemistry, followed by reduction to 1,2-diols. The primary alcohol was protected by the dimethoxytrityl group, and the remaining secondary alcohol converted to a phos-phitylating agent. These monomers could then be incorporated into oligomers under the standard conditions of automated DNA synthesis. The resulting molecules retain amino acid side-chains, but have the phosphodiester backbone more familiar to nucleic acids. 

The synthesis of two other important biopolymers, proteins and nucleic acids, also involves a sequence of functional group transformations. Simple (amidic or phosphodiester) bonds are formed between readily available monomeric units (amino acids or nucleotides). Almost all synthetic efforts in this area are centered around the elaboration of an optimal method to achieve an efficient formation of this bond. Given the complexity of the final structure, this task is never too simple. 

Figure 5.3. Backbones of DNA and RNA. The backbones of these nucleic acids are formed hy 3 -to-5 phosphodiester linkages. A sugar unit is highlighted in red and a phosphate group in blue.

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