Biological Chemistry - Berezov T. T., Korovkin B. F. 1998

Metabolism of Conjugated Proteins
Nucleic Acid Metabolism
Degradation of Nucleic Acids

Polymeric nucleic acid molecules are degraded in Tissues primarily via hydrolytic pathways involving specific Enzymes known as Nucleases. These are divided into endonucleases, which cleave internal internucleotide bonds in DNA and RNA molecules, causing nucleic acid depolymerization into oligonucleotides, and exonucleases, which catalyze the hydrolytic removal of terminal mononucleotides from DNA, RNA, or oligonucleotides. In addition to hydrolytic nucleases, there are enzymes that catalyze nucleic acid breakdown, for instance, via transferase reactions. They catalyze The transfer of a phosphoric acid residue from the 5'-carbon atom of the ribose of one mononucleotide to the 2'-carbon atom of an adjacent mononucleotide, accompanied by the Cleavage of the internucleotide bond and The formation of a phosphodiester bond between the 2'- and 3'-carbon atoms of the ribose of the same mononucleotide. To date, various groups of nucleases catalyzing the degradation of both DNA and RNA have been discovered.

Deoxyribonucleases I catalyze the cleavage of internal phosphodiester bonds in one of the two strands of a DNA molecule between the 3'-carbon atom of deoxyribose and the phosphate residue, yielding low-molecular-weight oligodeoxyribonucleotides:

Class="center">ДНК + (n-1) Н₂O -> n-Олигодезоксирибонуклеотиды.

The reaction products also include mono- and dinucleotides. Typical representatives of these enzymes are pancreatic DNases. One of them (DNase I) has been isolated in pure form, and The sequence of all its 257 amino acid residues has been elucidated. The enzyme exhibits maximum activity at pH 6.8–8.0, is activated by divalent Mg²⁺ and Mn²⁺ ions, and is inhibited by the End products of the enzymatic reaction, namely oligonucleotides.

Deoxyribonucleases II cause the depolymerization of DNA molecules via paired cleavages of the phosphodiester bonds in both DNA strands, yielding larger oligodeoxyribonucleotides. A representative of this group is Spleen-derived DNase II, which has a Molecular Weight of 38,000 and consists of 343 amino acid residues. Glucosamine has been identified as a component of this DNase. The enzyme is also activated by Metal Ions and inhibited by anions, with an optimal pH between 5.5 and 5.8.

In addition to these enzymes, exodeoxyribonucleases—which hydrolyze phosphodiester bonds in DNA molecules with the release of terminal 5'-deoxyribonucleotides—have been discovered (predominantly in microorganisms). For example, four such enzymes, designated as exodeoxyribonucleases I, II, III, and IV, have been isolated from E. coli.

Restriction Endonucleases (restriction enzymes), which operate via a DNase-like mechanism, catalyze The breakdown of foreign (primarily phage) DNA at strictly defined sites within palindromic sequences. Two such restriction enzymes, designated EcoRI and EcoRII, have been isolated from E. coli and characterized. Because restriction enzymes exhibit strict Specificity, they are utilized to decipher nucleotide sequences in phage and viral DNAs. Furthermore, this unique property of restriction enzymes is finding increasingly widespread Structure/182.html">Practical Application in Introduction/32.html">Genetic Engineering for "excising" specific DNA fragments and "integrating" them into bacterial DNA genomes to produce recombinant DNAs. As a result, Cells acquire a range of hereditary traits they did not previously possess. The theoretical and, above all, Practical significance of such research cannot be overstated. Evidence of the immense interest in genetic engineering is the establishment and successful Implementation of a joint comprehensive program—the "Restriction Enzymes" project—by institutes of the Russian Academy of Sciences and laboratories in several countries.

Many hundreds of restriction enzymes have been isolated in purified form and are now commercially available.

Among the enzymes that catalyze the hydrolytic breakdown of RNA, ribonucleases I have been studied the most thoroughly. They hydrolyze phosphodiester bonds within the RNA molecule. Isolated from the Pancreas of various animals, RNase consistently consists of 124 amino acid residues across species, although the enzymes show minor variations in Amino Acid Sequence; the Tertiary Structure of several RNases has also been elucidated (see Chapter 4). Guanylribonuclease, an enzyme that catalyzes the endonucleolytic cleavage of RNA, has been obtained in a homogeneous state from the mold fungus of the genus Aspergillus.

Among the enzymes that degrade DNA and RNA via non-hydrolytic pathways, polynucleotide phosphorylase and the DNA glycosylase group deserve mention. The physicochemical properties and Biological Role of microbial polynucleotide phosphorylase have been studied in detail in S.S. Debov's laboratory, where the enzyme was also discovered in animal tissues.

The MECHANISM OF ACTION of this enzyme involves the transfer of nucleotide residues from RNA to inorganic phosphate, yielding ribonucleoside diphosphate (RDP):

It is hypothesized that in vivo the enzyme catalyzes the breakdown of cellular RNAs, predominantly mRNA, into nucleoside diphosphates, thereby participating in the Regulation of cellular inorganic phosphate concentration. Another equally important and unique function of polynucleotide phosphorylase is its ability, in in vitro experiments, to catalyze the synthesis of polyribonucleotides with predefined sequences from free nucleoside diphosphates (NDPs). This enzyme played a pivotal role in deciphering METABOLISM/28.html">The Genetic Code in the laboratories of Nobel laureates S. Ochoa and M. Nirenberg (see Chapter 15).

A group of DNA glycosylases has been discovered that participate in the removal of modified purine and pyrimidine bases (e.g., uracil, which is formed upon the deamination of a cytosine residue in one of the DNA strands).

Thus, DNA glycosylases perform a crucial function in DNA Repair processes (restoration of the DNA Structure).

Through the sequential action of various cellular exo- and endonucleases, Nucleic Acids are degraded down to ribo- and deoxyribonucleoside 3'- and 5'-phosphates. The further breakdown of these products involves the enzymatic conversion of mononucleotides*, nucleosides, and subsequently free nitrogenous bases. At The First stage of Hydrolysis, 3'- and 5'-nucleotidases act to catalyze the hydrolytic breakdown of mononucleotides into free nucleosides, cleaving the inorganic phosphate from the C-3' or C-5' carbon atom of the carbohydrate moiety, respectively. At the Second Stage, the ribose residue is transferred from the nucleoside to free phosphoric acid, yielding ribose 1-phosphate and a free nitrogenous base.