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Nucleic acids are polymers of nucleotides. A nucleotide is a phosphate ester of a nucleotide. If is formed when a phosphate group is linked to OH of of a nucleoside through phosphoester linkage. Nucleosides are combinations of a base with a sugar. A nucleoside is an -glycoside formed between a base and OH of of a sugar (usually ribose or deoxyribose). The nucleotides found in cells are derivatives of heterocyclic, highly basic compounds.
Five major bases are found in cells. The derivatives of purine are called adenine and guanine, and the derivatives of pyrimidine are called thymine, cytosine, and uracil. The common abbreviations used for these five bases are A, G, T, C, and U.
The pyrimidines-thymine, uracil, and cytosine-are nitrogen heterocycles with a single ring. When conjugated to ribose, they are termed thymidine, uridine, and cytidine.
The purines-adenine and guanine-are nitrogen heterocycles with a double ring; their nucleosides are termed adenosine and guanosine.
Nucleosides are found in the cell primarily in their phosphorylated form. Nucleotides can exist in mono-, di-, or tri-phosphorylated forms. The nucleotide uridine is never found in DNA, and thymine is almost exclusively found in DNA. Thymine is found in tRNAs but not in rRNAs or mRNAs.
Nucleotide AND Nucleic Acid Nomenclature
· In 1868, Johann Friedrich Miescher developed a method for isolating the nuclei from white blood cells present in pus.
· The minute amounts of nuclear material that he obtained contained a novel substance that was slightly acidic and high in phosphorus. Miescher called this material nuclein, which consisted of DNA and protein.
· The substance was later renamed nucleic acid by one of his students, Altman.
· Phoebus Aaron Levene discovered that DNA consists of a large number of linked, repeating units called nucleotides.
E. Chargaff and his colleagues (1949) analysed the composition of DNA from various sources by quantitative chromatographic methods. Their observations are called Chargaff's rules:
· Regardless of the source, the purine and pyrimidine bases occur in equal amounts in a DNA molecule, i.e., .
· The amount of adenine is equivalent to the amount of thymine .
The amount of cytosine is equivalent to that of guanine . Thus, there is base equivalence in DNA.
The base ratio , however, is rarely equal to 1 and varies with different species from 0.4 to 1.9 (low in microorganisms and high in higher animals), but the ratio is constant for a species.
· Maurice Wilkins and Rosalind Franklin used X-ray diffraction to study DNA.
· They experimentally demonstrated that DNA was helical in structure.
· The correct three-dimensional structure of a DNA molecule was proposed in 1953 by James Watson and Francis Crick at Cambridge University.
· DNA exists as a helix of two complementary, antiparallel strands wound around each other in the rightward direction and stabilised by H -bonding between the bases in adjacent strands.
· The bases are in the interior of the helix aligned at nearly angle relative to the axis of the helix.
· Purine bases form hydrogen bonds with pyrimidines.
· From any fixed position in the helix, one strand is oriented in the direction and the other in the direction.
· Two chains are antiparallel.
· The bases of both strands lie on the inside of the structure and stack flat on one another.
· One complete turn of the helix is long and contains 10 bases.
· The double helix measures in diameter.
· The double helix of DNA has been shown to exist in several different forms depending on the sequence content and ionic conditions of crystal preparation.
· The B-form of DNA prevails under physiological conditions of low ionic strength and a high degree of hydration.
Nucleosomes are the fundamental organisational units of chromatin. DNA does not normally exist as a simple double helix. Instead, eukaryotic DNA is found packaged with protein, forming a substance called chromatin. Chromatin consists of fibres containing protein and DNA in approximately equal masses, along with a small amount of RNA.
The DNA in the chromatin is very tightly associated with proteins called histones, which package and order the DNA into structural units called nucleosomes. Also found in chromatin are many non-histone proteins, some of which help maintain chromosome structure, while others regulate the expression of specific genes.
Chromatin, therefore, consists of DNA wrapped around one histone octamer after another, like a long string of beads. The amount of DNA in a single human cell, lined up end to end, would stretch nearly 2 m .
· A nucleosome consists of a small amount of DNA wrapped up with protein. The proteins that interact with DNA to form chromatin comprise a family of basic (positively charged) proteins called histones.
· The disk-like nucleosome structure has a 10 nm diameter and a height of 5 nm .
· There are five different types of histone proteins: H1, H2A, H2B, H3, and H4.
Of these, two molecules each of H2A, H2B, H3, and H4 combine to form a histone octamer.
· Chromatin can be packed further into higher-order structures. This involves the action of histone H1.
· H1 binds to DNA on the outside of nucleosomes (at a ratio of one H 1 molecule per nucleosome). Then H 1 molecules interact with each other, causing the chromatin to form a spiral, with 6 to 8 nucleosomes per turn of the spiral. This structure is known as a solenoid or chromatin fibre. Because the chromatin is so tightly packed, DNA in the 30 nm fibre is genetically inactive.
· This configuration corresponds to heterochromatin or the state of chromosomes during cell division.
The 30 nm fibre is folded up further to make metaphase chromosomes.
Genetic material can transform bacteria. A British doctor S. F. Griffith first studied this transformation in 1928. He observed that Streptococcus pneumoniae, popularly known as Pneumococcus, has two strains: virulent and nonvirulent ( S and R , respectively).
· The virulent strain causes pneumonia. These bacteria are known as S-type because when grown on a suitable medium, they form smooth colonies.
· The nonvirulent type of bacteria does not produce the disease. They are, therefore, called rough or R-type (rough the small colonies).
Griffith could kill the bacteria by heating them. He observed that the heat-killed S-strain bacteria injected into mice did not kill the mice. When he injected a mixture of heat-killed bacteria and live bacteria, the mice died. Moreover, he recovered live bacteria from the carcasses.
He concluded that some substances needed to produce the mucous coat were passed from the dead bacteria (S strain) to the live ones ( strain), i.e. they became transformed.
Although Griffith did not understand the nature of transformation, he theorised that some substances in the polysaccharide coat of the dead bacteria might be responsible. He called this substance the transforming principle.
Prior to the work of Avery, MacLeod, and McCarty, the genetic material was thought to be protein. They worked to determine what the transforming substance was in Griffith's experiment. They purified chemicals from the heat-killed cells to see which ones could transform live cells into cells.
They discovered that DNA alone from S bacteria caused R bacteria to become transformed. They also discovered that protein-digesting enzymes (proteases) and RNA-digesting enzymes (RNAses) did not affect transformation, so the transforming substance was not a protein or RNA. Digestion with DNase did inhibit transformation, so DNA caused the transformation. They concluded that DNA was the hereditary material, but not all biologists were convinced.
The unequivocal proof that DNA is the genetic material came from the experiments of Alfred Hershey and Martha Chase (1952) on T2 bacteriophage. They grew some viruses on a medium that contained radioactive phosphorus and some others on a medium that contained radioactive sulphur. Hershey and Chase based their experiment on the fact that DNA, and not proteins, contains phosphorus, and similarly sulphur is present in proteins but not in DNA.
Viruses grown in the presence of radioactive phosphorus contained radioactive DNA but not radioactive protein because DNA contains phosphorus and protein does not. Similarly, viruses grown on radioactive sulphur contained radioactive protein but not radioactive DNA because DNA does not contain sulphur.
Radioactive phages were allowed to attach to Escherichia coli bacteria. Then, as the infection proceeded, the viral coats were removed from the bacteria by agitating them in a blender. The virus particles were separated from the bacteria by spinning them in a centrifuge.
Bacteria infected with viruses that had radioactive DNA were radioactive, indicating that DNA was the material that passed from the virus to the bacteria.
Bacteria infected with viruses that had radioactive proteins were not radioactive. This indicates that proteins did not enter the bacteria from the viruses.
DNA is, therefore, the genetic material that is passed from virus to bacteria.
Biology Faculty
M.Sc. Jiwaji University
