DNA

DNA 

  • DNA, or deoxyribonucleic acid, is indeed a lengthy polymer made up of deoxyribonucleotides. 
  • The length of DNA is typically characterized by the number of nucleotides or base pairs it contains, and this length is a unique trait of each organism. 
  • For instance, various organisms have different DNA lengths: the bacteriophage φ ×174 possesses 5386 nucleotides, the bacteriophage lambda has 48502 base pairs (bp), the bacterium Escherichia coli carries 4.6 × 10^6 bp of DNA, and the haploid content of human DNA comprises a staggering 3.3 × 10^9 bp. 

Salient features of the Double-Helix structure of DNA- 

  •      Comprises Two Polynucleotide Chains: DNA is composed of two polynucleotide chains. The backbone of these chains consists of alternating sugar-phosphate units, while the nitrogenous bases project inward. 

 

  • Anti-Parallel Polarity: The two chains run in opposite directions with anti-parallel polarity. If one chain has the 5' to 3' polarity, the other has the 3' to 5' polarity.

 

  •      Hydrogen Bonding and Base Pairs: The bases in the two strands are paired through hydrogen bonds (H-bonds), forming base pairs (bp). Adenine (A) pairs with thymine (T) through two H-bonds, and guanine (G) pairs with cytosine (C) through three H-bonds. This consistent pairing always places a purine opposite a pyrimidine, maintaining a uniform distance between the two strands. 
  •       Right-Handed Helix: The two DNA chains coil in a right-handed fashion. The helix has a pitch of approximately 3.4 nanometers (nm), with roughly 10 base pairs in each complete turn. This arrangement results in a distance of approximately 0.34 nm between adjacent base pairs. 

 

  •      Stacked Base Pairs: The plane of one base pair stacks directly over the plane of the adjacent base pair within the double helix. This stacking, along with H-bonds, contributes to the stability of the helical structure.

Central dogma 

The Central Dogma of Molecular Biology is a fundamental concept in genetics and molecular biology that describes the flow of genetic information within a biological system. Proposed by Francis Crick in 1958, it outlines the directional transfer of genetic information as follows: 

1. DNA Replication: The process begins with DNA replication, where a DNA molecule makes an exact copy of itself. This ensures the faithful transmission of genetic information from one generation of cells to the next during cell division.

 2. Transcription: The next step is transcription, in which a specific segment of DNA serves as a template to produce a complementary RNA molecule. This RNA molecule is known as messenger RNA (mRNA) and carries the genetic instructions encoded in DNA to the ribosome for protein synthesis. 

3. Translation: In translation, the information contained in the mRNA is used to synthesize proteins. Ribosomes read the mRNA's codons (three-base sequences) and match them to the appropriate amino acids, resulting in the assembly of a specific protein. 

The Central Dogma states that genetic information flows unidirectionally from DNA to RNA to protein. While this flow of information is considered a fundamental principle, it's important to note that there are exceptions, such as reverse transcription in retroviruses, where genetic information can flow from RNA back to DNA. However, the Central Dogma provides a foundational framework for understanding the core processes of gene expression in living organisms.

 

Packaging of DNA helix

  •  In a typical mammalian cell, if we calculate the length of the DNA double helix by multiplying the total number of base pairs (6.6 × 10^9 bp) with the distance between two consecutive base pairs (0.34 × 10^-9 m/bp), it amounts to approximately 2.2 meters. This length far exceeds the dimensions of a typical cell nucleus, which is around 10^-6 meters (1 micrometer). 
  •  In prokaryotes like Escherichia coli (E. coli), DNA is not scattered throughout the cell but is organized in a region called the 'nucleoid.' The DNA in the nucleoid is organized into large loops held by proteins. This helps in condensing and organizing the DNA within the relatively small cell. 
  • In eukaryotes, the organization is more complex. Positively charged, basic proteins called 'histones' play a crucial role. Histones have positively charged amino acid residues, primarily lysine and arginine, in their side chains. These positively charged histones form an octamer unit called a 'histone octamer.' The negatively charged DNA is wrapped around this histone octamer, forming a structure known as a 'nucleosome.' A typical nucleosome contains 200 base pairs of DNA helix. Nucleosomes are the repeating units of a structure in the nucleus called 'chromatin,' which appears as 'beads-on-string' when viewed under an electron microscope. 

 

 

  

  • Theoretical estimation of the number of nucleosomes (beads) present in a mammalian cell would depend on the size and DNA content of the cell but could be in the millions or more. 
  •      Chromatin fibers, composed of nucleosomes, are further coiled and condensed during the metaphase stage of cell division to form visible chromosomes. This higher-level packaging of chromatin requires additional proteins collectively referred to as 'Non-histone Chromosomal (NHC) proteins.' 
  •      Chromatin can exist in two forms: euchromatin and heterochromatin. Euchromatin is loosely packed and appears light when stained. It is considered transcriptionally active chromatin. In contrast, heterochromatin is densely packed and stains dark. It is considered inactive chromatin. 

Search for Genetic Material

  •     The quest to understand the mechanism of genetic inheritance began long ago, with significant contributions from scientists like Gregor Mendel, Walter Sutton, and Thomas Hunt Morgan. These early discoveries established that the genetic material was likely located within the chromosomes found in the nucleus of most cells. 
  •     In 1869, Friedrich Miescher discovered a substance he called "nuclein," which was later identified as DNA (deoxyribonucleic acid). This marked an important step in the search for the genetic material. 
  •      By 1926, scientists had made considerable progress in narrowing down their search for genetic material to the molecular level. However, the fundamental question of which molecule within the cell actually constituted the genetic material remained unanswered. 
  •      This search for the true nature of the genetic material was a complex and protracted scientific journey that involved various experiments, discoveries, and theories. It was a challenging puzzle to unravel. 
  •     It wasn't until the mid-20th century, specifically in 1953, that James Watson and Francis Crick proposed the now-famous double helix structure of DNA. This model provided strong evidence that DNA was the genetic material. 
  •      Additionally, the work of Rosalind Franklin and Maurice Wilkins, who generated X-ray diffraction data on DNA, was instrumental in elucidating its structure.
  •  Subsequent experiments and studies confirmed that DNA indeed carried the genetic information, providing a clear answer to the question that had puzzled scientists for decades. 
  •     The search for genetic material was a long and intricate scientific journey, ultimately leading to the discovery that DNA is the molecule responsible for carrying and transmitting genetic information in living organisms. This breakthrough revolutionized our understanding of genetics and laid the foundation for modern molecular biology.

 

Transforming Principle

 

  •      In 1928, Frederick Griffith conducted groundbreaking experiments involving Streptococcus pneumoniae, the bacterium responsible for pneumonia. His work revealed a phenomenon known as the "transforming principle." 
  •     Streptococcus pneumoniae bacteria can produce different types of colonies when grown on culture plates: smooth and shiny colonies (S strain) and rough colonies (R strain). The S strain bacteria possess a protective mucous (polysaccharide) coat, making them virulent, while the R strain lacks this coat and is non-virulent. 
  •      Griffith observed that heat-killing the S-strain bacteria rendered them non-virulent. When he injected these heat-killed S-strain bacteria into mice, the mice survived, indicating that the heat-killed S strain was harmless. 
  •      However, when Griffith injected a mixture of heat-killed S-strain bacteria and live R-strain bacteria into mice, the mice died from pneumonia. Remarkably, he also recovered living S-strain bacteria from the deceased mice.
  • Griffith's groundbreaking conclusion was that something from the heat-killed S strain bacteria, which he called the "transforming principle," had transformed the live R strain bacteria. This transformation allowed the R strain to synthesize a smooth polysaccharide coat, becoming virulent.

 

 

  •      Importantly, Griffith's experiments strongly suggested that the genetic material responsible for the transformation had been transferred. However, at this point, the biochemical nature of the genetic material had not been identified. 
  •      Griffith's work laid the foundation for the understanding of DNA as the genetic material, although the exact nature of the "transforming principle" would be further elucidated in subsequent experiments by other scientists, such as Avery, MacLeod, and McCarty in 1944. 

Biochemical characterization of the transforming principle by Oswald Avery, Colin MacLeod, and Maclyn McCarty:

  •      Prior to the work of Oswald Avery, Colin MacLeod, and Maclyn McCarty (1933-1944), the prevailing belief was that the genetic material was a protein. 
  •       Avery, MacLeod, and McCarty set out to determine the biochemical nature of the "transforming principle" in Griffith's experiment. They sought to identify whether it was protein, DNA, RNA, or another substance responsible for the transformation of R cells into S cells. 
  •      They conducted experiments where they purified various biochemical components (proteins, DNA, RNA, etc.) from the heat-killed S cells. They then individually mixed these components with live R cells to observe which ones could transform the R cells into S cells. 
  •      Astonishingly, they found that DNA alone from the S bacteria caused the transformation of R bacteria into S bacteria. This crucial discovery suggested that DNA was the genetic material responsible for the transformation.
  • To further confirm their findings, they conducted experiments with enzymes that could break down specific biomolecules. Protein-digesting enzymes (proteases) and RNA-digesting enzymes (RNases) were unable to prevent transformation. This indicated that the transforming substance was not a protein or RNA. 
  •      However, when they used DNase, an enzyme that specifically degrades DNA, they observed that it inhibited transformation. This provided strong evidence that DNA was indeed the hereditary material responsible for the transformation from non-virulent R cells to virulent S cells. 

 

 

  • Despite this groundbreaking work, not all biologists were initially convinced, and it took some time for the broader scientific community to fully accept that DNA was the genetic material. Nevertheless, Avery, MacLeod, and McCarty's research laid a solid foundation for our understanding of DNA as the molecule carrying genetic information, ultimately leading to the field of molecular genetics.

 

Hershey-Chase Experiment 

  •      Hershey and Chase in 1952 conducted experiments using bacteriophages, which are viruses that infect bacteria. Bacteriophages consist of genetic material (DNA or RNA) and a protein coat. 
  •      When a bacteriophage infects a bacterial cell, its genetic material enters the bacterial cell, and the bacterial cell treats this viral genetic material as if it were its own. The bacterial cell then replicates and produces more virus particles. 
  •      To determine whether it was protein or DNA from the viruses that entered the bacteria and served as the genetic material, Hershey and Chase devised a clever experiment. 
  •     They grew some bacteriophages in a medium containing radioactive phosphorus and others in a medium containing radioactive sulfur. Phosphorus is a component of DNA but not protein, while sulfur is a component of protein but not DNA. 
  •      Bacteriophages grown in the presence of radioactive phosphorus contained radioactive DNA but not radioactive protein because DNA contains phosphorus. 
  •      Conversely, bacteriophages grown in the presence of radioactive sulfur contained radioactive protein but not radioactive DNA because DNA does not contain sulfur. 
  •      In their experiment, they allowed these radioactive phages to attach to Escherichia coli (E. coli) bacteria. As the infection progressed, they agitated the bacteria in a blender to remove the viral coats and then separated the virus particles from the bacteria using a centrifuge. 
  •      Importantly, the bacteria infected with viruses containing radioactive DNA were found to be radioactive, indicating that DNA was the material passed from the virus to the bacteria. Conversely, bacteria infected with viruses containing radioactive proteins were not radioactive, providing evidence that proteins did not enter the bacteria from the viruses.

 

 

 

  •      These groundbreaking experiments conclusively demonstrated that DNA, and not protein, is the genetic material responsible for transmitting genetic information from viruses to bacteria. This was a pivotal moment in the history of molecular biology, solidifying the understanding that DNA carries the hereditary information in living organisms.

 

Properties of Genetic Material (DNA vs RNA) 

  •      The Hershey-Chase experiment provided definitive evidence that DNA is the genetic material. However, it's worth noting that some viruses, like Tobacco Mosaic viruses and QB bacteriophages, use RNA as their genetic material. 
  •      To understand why DNA is the predominant genetic material while RNA serves dynamic functions like messenger and adapter, we can examine two key chemical differences between DNA and RNA: 

1. Thymine vs. Uracil: DNA contains thymine (T) as one of its bases, while RNA contains uracil (U) in place of thymine. This substitution provides additional stability to DNA, making it less reactive and more chemically and structurally stable compared to RNA. 

2. 2'-OH Group: RNA has a 2'-OH (hydroxyl) group at every nucleotide, which makes it more reactive and easily degradable. In contrast, DNA lacks this 2'-OH group, rendering it less chemically reactive and structurally stable. 

  •      Genetic material must meet specific criteria, including the ability to replicate, stability, potential for slow changes (mutations), and the ability to express traits (Mendelian Characters). Both DNA and RNA can replicate due to base pairing and complementarity, whereas proteins cannot.
  • Stability is crucial for genetic material to remain unchanged throughout an organism's life cycle, age, or changes in physiology. DNA's stability is evident from Griffith's "transforming principle," where heat did not destroy its properties. The complementary nature of DNA strands allows them to reanneal after separation under appropriate conditions. RNA, with its 2'-OH group, is less stable and more labile.
  • Both DNA and RNA can undergo mutations, but RNA mutates at a faster rate due to its instability. RNA viruses, with shorter lifespans, mutate and evolve rapidly.
  • RNA can directly code for protein synthesis, making it capable of easily expressing traits. In contrast, DNA relies on RNA as an intermediary for protein synthesis, as the protein-synthesizing machinery has evolved around RNA.
  • While both RNA and DNA can function as genetic material, DNA is preferred for storing genetic information due to its greater stability. RNA, on the other hand, is better suited for the transmission and expression of genetic information. These distinctions highlight the complex and complementary roles of DNA and RNA in the genetic processes of living organisms.