(Unit II) Chapter 6: The Structure of DNA, and Its Importance For Holding Genetic Formation
I. Introduction
A. Introduction to DNA Structure: The Importance of DNA Structure
1. As we learned in Unit I, DNA is the Prime Genetic Molecule
a. Experiments By Hershey and Chase
b. Experiments by Avery Macleod and McCarty
2. This molecule must contain an incredible amount of information
3. Must contain information for proper development of an organism
a. Must allow the proper structures to form at the appropriate time
b. Must allow for appropriate growth at the appropriate time
4. Must contain the information for proper cellular function
a. Our DNA encodes proteins involved in respiration
b. Our DNA encodes proteins that are important in sending and receiving signals between cells
5. All the appropriate information is present to allow for reproduction
a. Cellular reproduction (asexual)
b. Organismal reproduction (sexual or asexual)
6. As Human Molecular Biologists we have a keen interest in the information in DNA as mistakes (Mutations) in DNA sequence lead to Genetic Disease
B. Introduction to DNA Structure: How It Holds The Information of Heredity
1. What is so special about DNA that gives it the ability to hold important genetic information?
2. The ability of DNA to hold all of this information lies in both its chemistry and 3-Dimensional structure
3. Several biochemists, including Friedrich Meisher were able to determine that DNA was composed of several components
a. Carbon
b. Phosphorous
c. Nitrogen
d. Hydrogen
4. Watson and Crick (1952) discovered that the 3-Dimensional structure of DNA is a double helix
a. Understood how the different atoms found in DNA are covalently linked together
b. Understood how those linkages are viewed in 3-Dimensions
5. The ability for DNA to hold genetic information lies in its structure
a. Compared to proteins, its structure seems quite simple
b. DNA has the ability to hold large amounts of stored information
6. From the 3-Dimensional structure of DNA came several implications
a. All genes would have roughly the same 3-Dimensional structure as most DNA in the cell takes the same 3-Dimensional shape
b. Differences between genes will come from the order and number of nucleotide building blocks
C. Introduction to DNA Structure: Differences in Structure Between DNA Molecules
1. Since Watson and Crick’s discovery, the basic principles of DNA chemistry and structure have not changed
2. However, each DNA molecule may not have the exact same structure
3. Some molecules are circular with no free ends, whereas other DNA molecules are linear and can have at least 2 free ends
4. Depending on how the DNA is crystallized, the double helix can take on any one of three forms
D. Introduction to DNA Structure: DNA is Not The Only Nucleic Acid
1. As a rule, nucleic acids are composed of carbon, nitrogen, phosphorous and hydrogen
2. DNA is not the only important nucleic acid found in nature
3. RNA is another type of nucleic acid also found in nature
a. For some viruses, this is their prime genetic molecule
b. In eukaryotic cells, RNA functions in expression of genes
c. In eukaryotic cells, RNA functions as a structural molecule for important intracellular components in the cell (ribosomes)
II. Types of Nucleic Acids: DNA and RNA
A. There are some fundamental similarities as well as differences between DNA and RNA
B. Below are some similarities
1. The basic units to make each are very similar
2. Each is polymerized in a very similar way
C. Below are some differences
1. DNA is double stranded, whereas RNA is single stranded
2. The basic building blocks (units) of RNA and DNA are slightly different (sugar and nitrogenous bases)
3. RNA is more chemically reactive
4. RNA can take more elaborate shapes
III. Building a DNA Molecule
A. Building the DNA Molecule: The Chemical Structure of Nucleic Acids
1. Nucleic Acids, whether DNA or RNA are polymers
2. DNA and RNA are composed of repeating units (monomers) called nucleotides
3. Each nucleotide consists of three basic components
a. Phosphate group
b. A five carbon sugar (deoxyribose)
c. A nitrogenous base
4. The phosphate group and the deoxyribose are part of the DNA backbone, whereas the nitrogenous bases are located towards the interior of the DNA molecule
B. Building the DNA Molecule: Nucleotide Structure and The Phosphate Group
1. The chemistry of the phosphate group is important in allowing DNA to be a polymer (i.e. the phosphate group is important in linking nucleotides together)
2. The phosphate group consists of a phosphorus and four oxygen atoms
3. The phosphorous is located centrally in the phosphate group, and each of the four oxygen atoms are bound to the phosphorous
4. The bonds between the phosphorous and each oxygen atom is unequal
a. They share electrons unequally
b. Oxygen atoms are slightly negative
c. Phosphorous is slightly positive
5. At physiological pH, the phosphate group is a proton donor
6. The linking bonds that are formed from the phosphates are esters
a. They have the property of being extremely stable
b. These bonds are easily broken by enzymatic hydrolysis (by adding water)
7. The fact that phosphate bonds are stable, yet easily broken is an important quality
a. Allows for polymerization of nucleotides
b. Allows for synthesis of DNA (or RNA) chains
C. Building the DNA Molecule: Nucleotide Structure and The Pentose Sugars
1. Each nucleotide will contain a pentose (5 carbon) sugar, whether it be a nucleotide that gets incorporated into DNA or RNA
2. The sugar that is used in DNA is deoxyribose, whereas ribose, is used in another type of nucleic acid called RNA
3. Within the ring, there are four carbon atoms (labeled 1’, 2’, 3’ etc) joined by an oxygen atom
4. The fifth carbon (the 5’ carbon) projects upward from the ring
D. Building the DNA Molecule: Nucleotide Structure and The Pentose Sugars
1. Ribose and Deoxyribose differ in structure only by the presence or absence of a 2’ hydroxyl group
a. For RNA, the 2’ carbon has a hydroxyl group bound to it
b. For DNA, the 2’ carbon does not have a hydroxyl group (deoxy) bound, instead it has a hydrogen bound to it
2. The difference in whether there is a 2’ hydroxyl (as in RNA) or a 2’ hydrogen (as in DNA), gives DNA and RNA different chemical properties due to the fact that the hydroxyl group is more reactive than the hydrogen
a. RNA can fold into a greater array of structures
b. DNA is more stable than RNA; RNA is more prone to degradation
E. Building the DNA Molecule: The Nitrogenous Base Component
1. The presence of the nitrogenous bases in nucleic acids was discovered by Friedrich Miecher after he started to determine the chemistry of his nuclein
2. They are called nitrogenous bases due to the fact that they are have a high nitrogen content
3. They are considered a base due to the fact that they have the properties of a base (proton acceptors)
4. By and large, when part of the structure of DNA the nitrogenous bases are non-polar, which is important for DNA structure
a. The bases are hydrophobic
b. The bases are located towards the interior of a molecule of DNA
5. There are five different types of nitrogenous bases in nucleic acids
a. Adenine
b. Guanine
c. Cytosine
d. Thymine
e. Uracil
6. Adenine and guanine are known as purines and have a double ring
7. Cytosine, Thymine and Uracil are known as pyrimidines and have a single ring
8. Both DNA and RNA contain Adenine, Guanine, Cytosine
9. Instead of Thymine as DNA has, RNA has Uracil
10. In nature, each nitrogenous base can take one of two conformations
11. Each base exists in two tautomeric states in equilibrium with each other
a. Tautomers are isomers that readily interconvert at equilibrium
b. Tautomerization results in the migration of a proton and a resulting shift from single to double bond, or vice versa
12. For the nitrogenous bases, there are two conformations
a. Conventional form
b. Tautomeric state
13. For all of the nitrogenous bases, the equilibrium strongly favors the conventional form
F. Building the DNA Molecule: Constructing a Nucleotide-The Basic Building Block of DNA
1. To build a nucleotide one must start with a nucleoside
2. A nucleoside consists of only a pentose sugar and a nitrogenous base
a. Guanosine (if containing guanine)
b. Cytosine (if containing cytidine)
c. Adenosine (if containing adenine)
d. Thymidine (if containing thymine)
e. Uridine (if containing uracil
3. A nucleoside becomes a nucleotide once at least one phosphate group is bound to the 5’ carbon)
a. If one phosphate is bound then it is a nucleotide monophosphate (NMP)
b. If two phosphates are bound then the nucleotide is a nucleotide diphosphate
c. If three phosphates are bound then the nucleotide is a nucleotide triphosphate
4. In a nucleotide, the nitrogenous base is bound to the 1’ carbon of the pentose sugar
5. In a nucleotide the phosphate group is bound to the 5’ carbon
6. Both the nitrogenous base and the phosphate group are added to the pentose via condensation reactions with water as the byproduct
7. A nucleotide can have one, two or three phosphates bound to the 5’ carbon
a. The phosphate that is bound to the 5’ carbon is known as the α phosphate
b. The second phosphate from the 5’ carbon is the β phosphate
c. The third phosphate from the 5’ carbon is the γ phosphate
H. Building the DNA Molecule: Naming the Nucleotides
1. To name a nucleoside, we use the name of the appropriate nitrogenous base in the nucleoside and add a sine, unless the nitrogenous base is thymine or uracil, then a dine is added
a. Guanosine (if containing guanine)
b. Cytosine (if containing cytidine)
c. Adenosine (if containing adenine)
d. Thymidine (if containing thymine)
e. Uridine (if containing uracil)
2. The name of a nucleotide comes uses as a root the name of the nucleoside followed by the number of phosphates the nucleotide contains
a. If one phosphate is bound then it is a nucleotide monophosphate (NMP)
b. If two phosphates are bound then the nucleotide is a nucleotide diphosphate
c. If three phophates are bound then the nucleotide is a nucleotide triphosphate
J. Building a DNA Molecule: A Strand of DNA Is Composed of Chains of Polynucleotides
1. To create a DNA strand, a polymer must be formed of repeating nucleotides
2. A strand of DNA is only formed in the 5’ à 3’ direction and never in the 3’ à 5’ direction
3. In forming a strand of DNA, the nucleotides will only be added onto the 3’ end of a growing DNA strand
4. In order to join two nucleotides together, a condensation reaction must occur between the free 3’OH group of the final nucleotide in a growing strand and the 5’ PO4 group in the nucleotide to be added
a. A phosphodiester bond is formed between the two nucleotides
b. A byproduct of the reaction is one molecule of water
K. Building the DNA Molecule: DNA Base Pairing
1. DNA is a double stranded molecule and therefore, two strands must be able to interact with each other
2. How the two strands interact were determined through 2 sets of experiments
a. Erwin Chargaff’s biochemical experiments
b. Watson and Crick’s X-ray diffraction studies
3. Chargaff wanted to determine the relative concentration of each nitrogenous base within a molecule of DNA
4. In 1940, Chargaff developed a paper chromatography method to analyze the amount of each nitrogenous base present in a molecule of DNA
5. Chargaff observed several important relationships among the molar concentrations of the different bases
6. In 1940 Chargaff proposed two important rules with regards to the nitrogenous base composition of DNA, which became known as Chargaff’s rules
7. Chargaff rules are as follows
a. [A] = [T]
b. [G] = [C]
c. [A] + [G] = [T] + [C] or the number of purines is equal to the number of pyrimidines
8. Chargaff also found that the base composition, as defined by the percentage of G and C (G+C content) for DNA is the basically the same for organisms of the same species, and different for organisms of different species
9. The G + C content can vary from 22 – 73% depending on the organism
10. Watson and Crick built off Chargaff’s work and determined that the secondary structure of DNA was a double helix
11. In the double helix, the two DNA strands interacted through base pairing
a. Adenine pairs with thymine (2 H bonds)
b. Guanine pairs with cytosine (3 H bonds)
12. To allow for proper base pairing, each strand in a DNA molecule must be anti-parallel
a. Allows the nitrogenous bases to align properly for efficient base pairing
b. The free 5’ ends of each strand are on opposite sides of the molecule
c. The free 3’ ends of each strand are also on opposite sides from each other
13. Base pairing is advantageous to the DNA chemistry
a. Excludes water from the interior of the DNA molecule