Molecular Biology PDF

# Basics of Molecular Biology ## Structure of Nucleic Acids (DNA and RNA) - Nucleic acids are the most important of all biomolecules. - They are found in abundance in all living things, where they function to create and encode and then store information in the nucleus of every living cell of eve...

# Basics of Molecular Biology ## Structure of Nucleic Acids (DNA and RNA) - Nucleic acids are the most important of all biomolecules. - They are found in abundance in all living things, where they function to create and encode and then store information in the nucleus of every living cell of every life-form organism on Earth. - The term nucleic acid is the overall name for DNA and RNA. They are composed of nucleotides, which are the monomers made of three components: - **5-carbon sugar (pentose)**: If the sugar is ribose, the nucleic acid is RNA (ribonucleic acid); if the sugar is deoxyribose, the nucleic acid is DNA (deoxyribonucleic acid). - **Phosphoric acid**: Phosphoric acid is attached to OH group at 3' carbon of deoxyribose sugar in one nucleotide and OH group at 5' carbon of deoxyribose sugar in adjacent nucleotide. - **Nitrogenous bases**: - They are organic molecules with a nitrogen atom that has the chemical properties of a base. - The main biological function of a nitrogenous base is to bond nucleic acids together. - Nitrogenous bases are typically classified as the derivatives of two parent compounds: pyrimidine and purine. - **Pyrimidines** are formed of 1 cyclic ring. Examples: Cytosine, thymine, uracil. - **Purines** are formed of 2 heterogeneous rings. Examples: Adenine, Guanine. ## There are five nitrogenous bases in total: - **Guanine**: Found in DNA and RNA. - **Adenine**: Found in DNA and RNA. - **Cytosine**: Found in DNA and RNA - **Thymine**: Found in DNA. - **Uracil**: Found in RNA | | | |-------------|-----------------------------------| | **Purines** | Double ring structures | | **Pyrimidines** | Single ring structures | **Figure 39** - Purines are present in both DNA and RNA. - As for pyrimidines, the distribution is different: DNA contains cytosine and thymine while RNA contains cytosine and uracil. Thus, in summary, DNA and RNA are polymers of nucleotides. - A nucleotide is composed of a nitrogenous base bound to a pentose sugar (both together form what is called **NUCLEOSIDE**), in addition to a phosphate group. - The number of phosphate groups can be either one group (nucleoside monophosphate), or 2 groups (nucleoside diphosphate), or 3 groups (nucleoside triphosphate). ## Formation of nucleotide polymers - Nucleotides are attached to each other through a 3’-5’ phosphodiester bond. - This bond links carbon number 3 of the sugar and carbon number 5 in the sugar of the next nucleotide. - Carbon number 5 links to this bond through sharing by its phosphate group. **Figure 42** - After polymerization is done, the formed chain will have a 3' end (in which carbon number 3 in the sugar is free and not involved in any bond) and a 5' end (in which carbon number 5 in the phosphate group of the sugar is free and not involved in the bond). ## Deoxyribonucleic acid (DNA) - DNA is located within the nucleus of the cell. - It is associated with basic proteins and forms chromosomes. This is **nuclear DNA**. - DNA is also located within mitochondrial matrix. This is **mitochondrial DNA (mtDNA)**. - The 2 strands of deoxyribonucleotides wind around each other in a clockwise direction, forming a structure called **double helix**. - In the double helix, the sugar and phosphates are located to the exterior, forming the backbone, while the nitrogenous bases are located to the interior for better protection from degradation. ## Properties of DNA strands - Each DNA strand has polarity. - **3' End (3 prime)**: It is the end of DNA strand where C3 of deoxyribose sugar is not linked and is free. - **5' End (5 prime)**: It is the end of DNA strand where C5 of deoxyribose sugar is not linked and is free. - Therefore, two DNA strands show polarity, where each strand shows polarity either in 3'-5' direction or 5'-3' direction. - In DNA helix, two strands run in antiparallel direction; one strand runs from 3' -5' direction while the other runs from 5'-3' direction. - **Base Pairing**: Base pair signifies two nitrogenous bases held together through hydrogen bonding. Each pair of base consists of purine in one strand and pyrimidine in another strand. Particular purine always pairs with particular pyrimidine. - **Complementary base pairing**: A=T, G=C ## Functions of DNA 1. DNA has a unique property of duplicating itself into an exact copy. It is essential for transmission of characters from parents to offspring. Hence, it is a genetic material of cells. 2. DNA can transcribe mRNA on its strand. The mRNA contains codons for protein synthesis. 3. DNA regulates synthesis of proteins through codons. 4. DNA controls gene expression. It can regulate cellular functions like secretion, excretion, proliferation, and apoptosis. 5. DNA can undergo mutations which are essential for origin of species. 6. It controls differentiation of cells in embryonic development. ## Ribonucleic acid (RNA) - RNA is present mainly in the cytoplasm of the cell and has three types: ### 1. Messenger RNA (mRNA): - It constitutes 5% of all RNA. - It is synthesized in the nucleus by DNA and then sent to the ribosomes in the cytoplasm. - The mRNA carries a message from DNA in the nucleus, where it is synthesized, to the ribosomes in the cytoplasm, where it is going to direct the synthesis of a specific protein. - The letters of the message are the nitrogenous bases, the sequence of which is responsible for arranging the amino acids in proper order in the polypeptide chain to be synthesized. - Each 3 successive bases in the mRNA are called a "codon" because they code for a specific amino acid. **Figure 45** - The process of synthesis of mRNA in the nucleus under the directions of DNA is called "transcription". ### 2. Transfer RNA (tRNA): - It constitutes 15% of all RNA. - It is present in the cytoplasm and may be also known as soluble RNA (sRNA). - It is composed of 74 to 95 nucleotides. - tRNA molecules function as carriers for the amino acids and transfer them to the machinery site of protein synthesis in the cell (Ribosome). - There is, at least, a specific tRNA for each amino acid. - This means that there are least, 20 different tRNA molecules in every cell, each of which is specialized to carry one of the 20 different amino acids required for the process of protein synthesis. **Figure 46** ### 3. Ribosomal RNA (rRNA): - It constitutes 80% of total RNA. - It is present in the ribosomes of the cytoplasm. - The ribosome is a cytoplasmic nucleoprotein structure which acts as the machinery for the synthesis of proteins. - On the ribosomes, the mRNA and tRNA molecules interact to translate a specific protein. - The ribosome particle is formed of at least four rRNA molecules complexed with several protein molecules. - The ribosomal particle consists of 2 subunits. - The larger subunit is known as 60S, while - the smaller one is known as 40S (S= Svedberg unit, measured by ultracentrifugation). **Figure 47** ## Comparison between DNA and RNA | Points of comparison | DNA | RNA | |-----------------------|------------------------------------------|--------------------------------------------| | Occurrence | Chiefly in the nucleus | Chiefly in the cytoplasm | | Shape | Double helix | Single strand | | Types | One type | Three types, mRNA, tRNA, and rRNA | | Sugar present | Deoxyribose | Ribose | | Bases: Purines | Adenine, guanine, cytosine, thymine | Adenine, guanine, cytosine, uracil | | Function | Form genes and direct synthesis of mRNA, tRNA, rRNA | Protein synthesis | ## Replication of DNA - Replication of DNA is the biological process of synthesizing two DNA molecules from a parental DNA molecule. - In Eukaryotes, DNA replication occurs in the nucleus during S-phase of interphase (interval between two cell divisions). ## Rules of DNA replication in eukaryotes: 1. **DNA replication is semiconservative:** Each DNA strand serves as a template for synthesis of a new strand producing two DNA molecules, each with one new strand and one old strand. **Figure 48** 2. **Replication begins at multiple origins and usually proceeds bidirectionally:** Having multiple origins of replication provides a mechanism for rapidly replicating the great length of eukaryotic DNA molecules. **Figure 49** 3. **Replication exhibits polarity:** DNA synthesis proceeds in a 5` 3` direction and is semidiscontinuous. 4. **Replication is very accurate:** Replication proceeds with an extraordinary degree of fidelity. 5. **DNA replication occurs in the nucleus during the synthetic (S) phase of the eukaryotic cell cycle.** This phase is preceded and followed by two periods during which DNA is not synthesized (gap periods G1, and G2). During cell division (mitotic; M phase), each daughter cell receives one of the two identical DNA molecules. 6. **DNA replication is semi-discontinuous.** One strand (the leading strand) is synthesized in a continuous manner. The other strand (the lagging strand), is synthesized in the form of short fragments (Okazaki fragments), that are joined together to form one strand after polymerization is over. **Figure 50** ## Prerequisites of DNA Replication 1. **Deoxyribonucleoside Monophosphates (dAMP, dGMP, dTMP, and dCMP)**: Deoxyribonucleoside monophosphates constitute raw material for DNA replication. ## Enzymes Involved in DNA Replication 1. **DNA helicase**: Helicase moves along the sugar-phosphate backbone and disrupts hydrogen bonds in annealed nucleotide bases (complementary bases held by hydrogen bonds). Helicase's function is to separate helically coiled DNA strands. 2. **DNA topoisomerase**: It induces break in DNA strand to relieve coiling stress. 3. **DNA ligase**: It is helpful in DNA replication and repair. It helps to join two Okazaki fragments after removal of RNA primer. It seals DNA segments after correction of mismatched nucleotide bases. 4. **Primase**: It is a type of RNA polymerase. It catalyzes formation of RNA primer on DNA template. 5. **Telomeres**: They are repetitive sequences TTAGGGs, these telomeres (Hexamers) cap end of human chromosomes and are believed to prevent chromosomes from undergoing degradation. Telomeres are believed to contribute to the preservation of the normal aging process. With each round of replication, the length of telomeres is shortened. During normal cell division, the length of telomeres is shortened, thus limiting the life span of the cell. Therefore, normal somatic cells after certain number of cell division die d to shortening of telomeres. So telomeres determine the proliferation capacity of human somatic cells. ## N.B. DNA is degraded by nucleases or DNases: - They may be either exonucleases or endonucleases. - **Exonucleases**: Degrade nucleic acids from one end of the molecule operating either from 5' 3' or from 3' 5' direction of one strand of the double strand nucleic acids. - **Endonucleases**: Can begin to degrade at any internal site in a nucleic acid strand, reducing it to smaller and smaller fragments.