Biochemistry and Molecular Biology - Belyasova N.A. 2002

Molecular Foundations and Mechanisms of Heredity
Organization of the Cellular Genetic Apparatus
Types of Nucleic Acids and Their Functions

Of the Two Types of Nucleic Acids—DNA and RNA—deoxyribonucleic acid serves as the molecule that encodes all the essential hereditary information of The Cell and is capable of self-Replication, whereas Ribonucleic Acids act as intermediaries between DNA and Proteins. These Functions of nucleic acids are closely related to the specifics of their individual Structure.

DNA and RNA are polymeric macromolecules whose monomers are NUCLEOTIDES. Each nucleotide consists of three parts: a monosaccharide, a phosphoric acid residue, and a nitrogenous base. The nitrogenous base is linked to the sugar via a ß-N-glycosidic bond (Fig. 1.1).

The sugar component of the nucleotide (a pentose) can exist in one of two forms: ß-D-ribose or ß-D-2-deoxyribose. The difference between them is that the ribose hydroxyl group at the 2'-carbon atom of the pentose is replaced by a hydrogen atom in deoxyribose. Nucleotides containing ribose are called ribonucleotides and serve as the monomers of RNA, whereas nucleotides containing deoxyribose are termed deoxyribonucleotides and form DNA.

Nitrogenous bases are derivatives of one of two compounds: purine or pyrimidine. Nucleic acids predominantly contain two purine bases—adenine (A) and guanine (G)—and three pyrimidine bases—cytosine (C), thymine (T), and uracil (U). Ribonucleotides (and consequently RNA) contain A, G, C, and U, whereas deoxyribonucleotides (and DNA) contain A, G, C, and T.

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Fig. 1.1. Structure of a nucleoside and a nucleotide: the numbers indicate the positions of atoms in the pentose residue

The nomenclature of Nucleosides and Nucleotides is widely used in biochemistry and molecular biology and is presented in Table 1.1.

Table 1.1. Nomenclature of nucleotides and nucleosides

Long polynucleotide chains of DNA and RNA are formed by linking nucleotides together through phosphodiester bonds. Each phosphate connects the hydroxyl group at the 3'-carbon atom of one nucleotide's pentose to the OH group at the 5'-carbon atom of the adjacent nucleotide's pentose (Fig. 1.2).

Acid Hydrolysis of nucleic acids yields individual nucleotide components, whereas Enzymatic hydrolysis using Nucleases cleaves specific bonds within the phosphodiester backbone, thereby exposing the 3'- and 5'-ends of the molecule (Fig. 1.2).

Fig. 1.2. Introduction/11.html">Secondary structure of DNA. Enzymatic hydrolysis of the DNA chain exposing the 3'- and 5'-free ends of the molecule

This provides a basis for considering the nucleic acid chain to be polarized, making it possible to define the direction for reading The nucleotide sequence. It should be noted that most Enzymes involved in the synthesis and hydrolysis of nucleic acids operate in the 5'-to-3' direction (5' → 3') of the nucleic acid chain. By convention, the nucleotide sequence in nucleic acid chains is also read in the 5' → 3' direction (Fig. 1.2).

Structural Features of DNA. According to the three-dimensional model proposed by Watson and Crick in 1953, the DNA molecule consists of two polynucleotide chains that form a right-handed double helix around a common axis. The directions of the two chains are antiparallel, and the molecule maintains a nearly constant diameter and other parameters independent of its nucleotide composition, unlike proteins, in which the Amino Acid Sequence determines the secondary and Tertiary Structure of the molecule.

The sugar-phosphate backbone is located on the periphery of the helix, while the nitrogenous bases are on the inside, with their planes perpendicular to the helical axis. Specific Hydrogen Bonds form between bases facing each other in opposite chains: adenine always pairs with thymine, and guanine with guanine—correction: guanine with cytosine. Furthermore, in an AT pair, the bases are joined by two hydrogen bonds: one forms between the amino and keto groups, and the other between two nitrogen atoms of the purine and pyrimidine, respectively. In a GC pair, there are three hydrogen bonds: two form between the amino and keto groups of the respective bases, and the third forms between the nitrogen atom of the pyrimidine and the hydrogen (attached to the nitrogen atom) of the purine.

Thus, the bulkier Purines always pair with the smaller Pyrimidines. As a result, the distances between the C1' atoms of deoxyribose in the two chains are identical for both AT and GC pairs, measuring 1.085 nm. These two types of nucleotide pairs, AT and GC, are referred to as complementary pairs. Pairing between two purines, two pyrimidines, or non-complementary bases (A+C or G+T) is sterically hindered because appropriate hydrogen bonds cannot form, thereby disrupting the geometry of the helix.

The geometry of The Double Helix is such that adjacent nucleotides in the chain are spaced 0.34 nm apart. There are 10 nucleotide pairs per turn of the helix, and the pitch of the helix is 3.4 nm (10 x 0.34 nm). The diameter of the double helix is approximately 2.0 nm. Because the sugar-phosphate backbone is located further from the helical axis than the nitrogenous bases, the double helix features grooves—a major groove and a minor groove (Fig. 1.3).

The DNA molecule can adopt various Conformations, with A-, B-, and Z-forms having been identified. B-DNA is the conventional form in which DNA exists within the cell; in this form, the planes of the base rings are perpendicular to the axis of the double helix. In the A-form of DNA, the planes of the Base Pairs are tilted by approximately 20° relative to the normal to the axis of the right-handed double helix. Z-DNA is a left-handed helix with 12 nucleotide pairs per turn. The BIOLOGICAL FUNCTIONS OF the A- and Z-forms of DNA are not yet fully understood.

Fig. 1.3. Schematic representation of the B-form of the DNA double helix, showing the Major and minor grooves. The distance between adjacent base pairs and the pitch of the helix are indicated [M. Singer, P. Berg, 1998]

The Stability of the double helix is maintained by hydrogen bonds between complementary nucleotides in the antiparallel strands, stacking interactions (interplanar Van der Waals contacts between atoms and overlap of π-orbitals of the contacting bases), and hydrophobic interactions. The latter are manifested by the nonpolar nitrogenous bases being tucked into the interior of the helix, shielded from direct contact with the polar solvent, whereas the charged sugar-phosphate groups face outward and interact with the solvent.

Because the two DNA strands are held together solely by noncovalent bonds, the DNA molecule readily dissociates into single strands upon heating or in alkaline solutions (Denaturation). However, upon slow cooling (annealing), the strands can re-associate, and Hydrogen bonds are restored between complementary bases (renaturation). These properties of DNA are of great importance for Genetic Engineering methodology (Chapter 20).

The size of DNA molecules is expressed in terms of the number of nucleotide pairs, with one thousand base pairs (kbp) or 1 kilobase (kb) serving as the unit. The molecular mass of 1 kbp of B-DNA is ~ 6.6⋅10⁵ Da, and its length is 340 nm. The complete genome of E. coli (~ 4⋅10⁶ bp) is represented by a single circular DNA molecule (the nucleoid) with a length of 1.4 mm.

Structural features and functions of RNA. RNA molecules are single-stranded polynucleotides consisting of 70 to 10,000 nucleotides (sometimes more), represented by the following types: mRNA (Messenger RNA), tRNA (Transfer RNA), rRNA (ribosomal RNA), and—exclusively in Eukaryotic Cells—hnRNA (heterogeneous nuclear RNA) and snRNA (small nuclear RNA). These types of RNA perform specific functions; moreover, in some Viral Particles, RNA serves as the carrier of Genetic information.

Messenger RNA is a transcript of a specific segment of the DNA sense strand and is synthesized during METABOLISM/31.html">Transcription. mRNA serves as the blueprint (template) used to construct a polypeptide molecule. Every three consecutive nucleotides in mRNA function as a codon, determining THE POSITION OF the corresponding amino acid within the peptide. Thus, mRNA acts as an intermediary between DNA and protein.

Transfer RNA also plays a vital role in Protein Synthesis. Its function is to deliver Amino Acids to the site of synthesis and determine their placement within the peptide chain. To achieve this, tRNA features a specific triplet of nucleotides known as an "anticodon," and the molecule as a whole possesses a unique structure. The structural model of a tRNA molecule is traditionally referred to as a "cloverleaf" (Fig. 1.4).

The tRNA molecule is relatively short, consisting of 74–90 nucleotides. Like any nucleic acid chain, it has two distinct ends: a phosphorylated 5' end and a 3' end, which always terminates with the CCA nucleotide sequence and a 3'-OH group. The amino acid attaches to the 3' end, which is why it is called the acceptor end. tRNA molecules also contain several unusually modified nucleotides that are not found in Other types of nucleic acids.

Fig. 1.4. Structure of tRNA [E. Rees, M. Sternberg, 1988]

Although the tRNA molecule is single-stranded, it contains localized duplex regions that form so-called stems or arms, where Watson—Crick base pairs form between asymmetric sections of the chain (Fig. 1.4). All known tRNAs form a cloverleaf structure with four stems (the acceptor, D, anticodon, and T stems). These stems adopt the conformation of a right-handed double helix, known as A-form DNA. The loops of tRNA represent single-stranded regions. Some tRNAs possess additional loops and/or stems (for example, the variable loop in Yeast phenylalanine tRNA).

Recognition of the corresponding site on the mRNA by the tRNA molecule is mediated by the anticodon located in the anticodon loop (Fig. 1.4). This process involves The formation of hydrogen bonds between the codon and anticodon bases, provided that their constituent sequences are complementary and the polynucleotide chains are antiparallel (Fig. 1.5).

While different tRNA molecules vary in their nucleotide sequences, their tertiary structures are remarkably similar. The molecule is folded in such a way that it resembles the letter L in shape. The acceptor and T-stems are specially arranged in space to form a continuous helix, constituting the "crossbar" of the L, while the anticodon and D-stems form the "stem." Proper spatial folding of tRNA molecules is essential for their biological function.

Quantitatively, ribosomal RNA is the most abundant RNA in the cell; however, its diversity compared to other RNA types is the lowest. rRNA accounts for up to 80% of total cellular RNA mass and is represented by only three to four distinct species. Meanwhile, the mass of nearly 100 different tRNA species accounts for about 15%, whereas thousands of various mRNAs make up less than 5% of cellular RNA mass.

E. coli cells contain Three types of rRNA: 5 S, 16 S, and 23 S, whereas eukaryotic cells utilize 18 S, 5.8 S, 28 S, and 5 S rRNAs. These species are integral components of Ribosomes, making up approximately 65% of their mass. Within ribosomes, rRNAs are tightly packed and capable of folding to form base-paired stems similar to those found in tRNAs. It is believed that rRNAs participate in binding the ribosome to tRNA; specifically, studies have shown that 5 S rRNA interacts with the T-arm of tRNA.

In addition to the aforementioned RNA types, eukaryotic nuclei contain heterogeneous nuclear RNA (hnRNA) and small nuclear RNA (snRNA). hnRNA accounts for less than 2% of total cellular RNA. These molecules undergo rapid turnover, with most having a half-life of no more than 10 minutes. One of the few established functions of hnRNA is its role as an mRNA precursor. snRNAs are associated with various proteins to form small nuclear ribonucleoprotein particles (snRNPs), which carry out RNA splicing (Chapter 3).

Fig. 1.5. Interaction between the mRNA codon and the tRNA anticodon. Dots indicate hydrogen bonds between complementary nucleotides