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Lecture 1 (Chapter 9)
MOLECULAR STRUCTURE OF DNA AND RNA
DNA is the Genetic Material
Genetics: Study of the structure, function, transmission of genes
Only living organisms have genes To understand genetics, start with the question:
“What is Life?” Characteristics shared by all living forms but not by
non-living forms
DNA is the Genetic Material DNA carries information Information, entropy and Solla Sollew
“Duhka (A wheel out of kilter) I Had Trouble in Getting to Solla Sollew (where there aren’t any
problems at least very few)
LIFE (GENES)
(negative entropy)
UNIVERSE
(entropy)
DNA is the Genetic Material To create and maintain information and order,
most living organisms “suck energy” (directly or indirectly) from the Sun
Short mean λ = high radiation energy, low entropy
If life, long λ = low radiation energy high entropy
If no life, medium λ = medium radiation energy, medium
entropy
Very high potential energy;
Very low entropy
Fusion reactions in sun’s core Hydrogen to helium with release of photons
Mass converted to energy (E = mc2) 600 million tons hydrogen fused to helium per second Creates 596 tons helium per second + 3.9 x 1026 Joules of energy Life borrows this energy to build complex, highly ordered structures
DNA is the Genetic Material The source of all life’s “problems” is entropy The “solution” is stored as information in DNA
Information and negative entropy Analog vs digital
Different organisms have different genes; different strategies for the survival of the genes and their transmission to the next generation
Those genes that built the best “survival machines” are still with us today
The vast majority of genes were lost
Basic Properties of DNA To fulfill its role, the genetic material must meet
several criteria: 1. Information: It must contain the information necessary
to make an entire organism (often from a single-celled zygote!)
2. Transmission: It must be passed from parent to offspring
3. Replication: It must be copied In order to be passed from parent to offspring
4. Variation: It must be capable of changes To evolve
Identification of DNA as Life’s Information Molecule
The identification of DNA as the genetic material involved a series of outstanding experimental approaches Griffith (1928) Avery, MacLeod, and McCarty (1944) Hershey and Chase (1952)
Griffith studied a bacterium (Diplococcus pneumoniae) now known as Streptococcus pneumoniae
S. pneumoniae comes in two strains S Smooth
Secrete a polysaccharide capsule Protects bacterium from the immune system of animals so is
virulent (i.e. deadly) Produce smooth colonies on solid media
R Rough Unable to secrete a capsule
Destroyed by animals’ immune system so is non-virulent Produce colonies with a rough appearance
Frederick Griffith’s Experiments with Streptococcus pneumoniae
Figure 9.2
Griffith concluded that something from the dead type IIIS was transforming type IIR into type IIIS He called this process transformation Whatever was “transforming” the IIR bacteria was not
sensitive to heat
Griffith called this the transformation principle Griffith did not know what it was
The chemical nature of the transforming principle was determined using experimental approaches that incorporated various biochemical techniques
Avery, MacLeod and McCarty realized that Griffith’s observations could be used to identify the genetic material
They carried out their experiments in the 1940s At that time, it was known that DNA, RNA, proteins and
carbohydrates are major constituents of living cells They prepared cell extracts from type IIIS cells
containing each of these macromolecules Only the extract that contained purified DNA was able
to convert type IIR into type IIIS
The Experiments of Avery, MacLeod and McCarty
In 1952, Alfred Hershey and Marsha Chase provided further evidence that DNA is the genetic material
Hershey and Chase Experiment with Bacteriophage T2
They studied the bacteriophage T2 It is relatively simple
since its composed of only two macromolecules
DNA and protein
Figure 9.4
Made up of protein
Inside the capsid
Figure 9.5
Life cycle of the T2 bacteriophage
The Hershey and Chase experiment can be summarized as follows: Used radioisotopes to distinguish DNA from proteins
32P labels DNA specifically 35S labels protein specifically
Radioactively-labeled phages were used to infect nonradioactive Escherichia coli cells
After allowing sufficient time for infection to proceed, the residual phage particles were sheared off the cells
=> Phage ghosts and E. coli cells were separated Radioactivity was monitored using a scintillation
counter
Figure 9.6
Figure 9.6
Interpreting the Data Most of the 35S was found in the
supernatantBut only a small percentage of 32P
These results suggest that DNA is injected into the bacterial cytoplasm during infection
This is the expected result if DNA is the genetic material
In 1956, A. Gierer and G. Schramm isolated RNA from the tobacco mosaic virus (TMV), a plant virus Purified RNA caused the same lesions as intact TMV
viruses Therefore, the viral genome is composed of RNA
Since that time, many RNA viruses have been found Refer to Table 9.1
RNA Functions as the Genetic Material in Some Viruses
DNA and RNA are large macromolecules with several levels of complexity
1. Nucleotides form the repeating units 2. Nucleotides are linked to form a strand 3. Two strands can interact to form a double helix 4. The double helix folds, bends and interacts with
proteins resulting in 3-D structures in the form of chromosomes
Nucleic Acid Structure
Copyright ©The McGraw-Hill Companies, Inc. Permission required for reproduction or display
Three-dimensional structureFigure 9.7
The nucleotide is the repeating structural unit of DNA and RNA
It has three components A phosphate group A pentose sugar A nitrogenous base
Refer to Figure 9.8
Nucleotides
Figure 9.8
Figure 9.9 The structure of nucleotides found in (a) DNA and (b) RNA
A, G, C or T
These atoms are found within individual nucleotides However, they are removed when nucleotides join together to make
strands of DNA or RNA
A, G, C or U
Base + sugar nucleoside Example
Adenine + ribose = Adenosine Adenine + deoxyribose = Deoxyadenosine
Base + sugar + phosphate(s) nucleotide Example
Adenosine monophosphate (AMP) Adenosine diphosphate (ADP) Adenosine triphosphate (ATP)
Refer to Figure 9.10
Figure 9.10
Base always attached here
Phosphates are attached there
Nucleotides are covalently linked together by phosphodiester bonds A phosphate connects the 5’ carbon of one nucleotide to
the 3’ carbon of another Therefore the strand has directionality
5’ to 3’
The phosphates and sugar molecules form the backbone of the nucleic acid strand The bases project from the backbone
Figure 9.11
In 1953, James Watson and Francis Crick discovered the double helical structure of DNA
The scientific framework for their breakthrough was provided by other scientists including Linus Pauling Rosalind Franklin and Maurice Wilkins Erwin Chargaff
A Few Key Events Led to the Discovery of the Structure of DNA
In the early 1950s, he proposed that regions of protein can fold into a secondary structure -helix
Linus Pauling
Figure 9.12
To elucidate this structure, he built ball-and-stick models Refer to Figure 9.12b
She worked in the same laboratory as Maurice Wilkins
She used X-ray diffraction to study wet fibers of DNA
Rosalind Franklin
The diffraction pattern is interpreted (using mathematical theory)
This can ultimately provide information concerning the
structure of the molecule
She made marked advances in X-ray diffraction techniques with DNA
The diffraction pattern she obtained suggested several structural features of DNA
Helical More than one strand 10 base pairs per complete turn
Died of ovarian cancer at the age of 38 Would have shared the Nobel Prize with Wilkins, Watson, and
Crick had she lived long enough (1962) Cancer may have resulted from exposure to X-rays
Rosalind Franklin
Chargaff pioneered many of the biochemical techniques for the isolation, purification and measurement of nucleic acids from living cells
It was already known then that DNA contained the four bases: A, G, C and T
Erwin Chargaff’s Experiment
The Hypothesis An analysis of the base composition of DNA in
different species may reveal important features about the structure of DNA
Testing the Hypothesis Refer to Figure 9.14
Figure 9.14
Figure 9.14
The Data
Interpreting the Data
The data shown in Figure 9.14 are only a small sampling of Chargaff’s results
The compelling observation was that Percent of adenine = percent of thymine Percent of cytosine = percent of guanine
This observation became known as Chargaff’s rule It was crucial evidence that Watson and Crick used to
elucidate the structure of DNA
Familiar with all of these key observations, Watson and Crick set out to solve the structure of DNA They tried to build ball-and-stick models that incorporated
all known experimental observations
A critical question was how the two (or more strands) would interact An early hypothesis proposed that the strands interact
through phosphate-Mg++ crosslinks Refer to Figure 9.15
Watson and Crick
Figure 9.15
This hypothesis was, of course, incorrect!
They went back to the ball-and-stick units They then built models with the
Sugar-phosphate backbone on the outside Bases projecting toward each other
They first considered a structure in which bases form H bonds with identical bases in the opposite strand ie., A to A, T to T, C to C, and G to G
Model building revealed that this also was incorrect
Watson and Crick
They then realized that the hydrogen bonding of A and T resembled that between C and G So they built ball-and-stick models with AT and CG
interactions These were consistent with all known data about DNA
structure Refer to Figure 9.16
Watson, Crick, and Maurice Wilkins were awarded the Nobel Prize in 1962
Watson and Crick
General structural features (Figures 9.17 & 9.18)
The DNA Double Helix
Two strands are twisted together around a common axis
There are 10 bases per complete twist The two strands are antiparallel
One runs in the 5’ to 3’ direction and the other 3’ to 5’ The helix is right-handed
As it spirals away from you, the helix turns in a clockwise direction
General structural features (Figures 9.17 & 9.18)
The DNA Double Helix
The double-bonded structure is stabilized by
1. Hydrogen bonding between complementary bases A bonded to T by two hydrogen bonds C bonded to G by three hydrogen bonds
2. Base stacking Within the DNA, the bases are oriented so that the flattened
regions are facing each other
General structural features (Figures 9.17 & 9.18)
The DNA Double Helix
There are two asymmetrical grooves on the outside of the helix
1. Major groove
2. Minor groove
Certain proteins can bind within these grooves They can thus interact with a particular sequence of bases
Figure 9.17
Figure 9.18
To fit within a living cell, the DNA double helix must be extensively compacted into a 3-D conformation This is aided by DNA-binding proteins
Refer to 9.21
The Three-Dimensional Structure of DNA
Figure 9.21
DNA wound around histone
proteins
The primary structure of an RNA strand is much like that of a DNA strand Refer to Figure 9.22 vs. 9.11
RNA strands are typically several hundred to several thousand nucleotides in length
In RNA synthesis, only one of the two strands of DNA is used as a template
RNA Structure
Figure 9.22
Although usually single-stranded, RNA molecules can form short double-stranded regions This secondary structure is due to complementary base-
pairing A to U and C to G
This allows short regions to form a double helix
RNA double helices typically Are right-handed Have 11 to 12 base pairs per turn
Different types of RNA secondary structures are possible Refer to Figure 9.23
Figure 9.23
Complementary regions
Noncomplementary regions
Held together by hydrogen bonds
Have bases projecting away from double stranded regions
Many factors contribute to the tertiary structure of RNA For example
Base-pairing and base stacking within the RNA itself
Interactions with ions, small molecules and large proteins
Figure 9.24 depicts the tertiary structure of tRNAphe
The transfer RNA that carries phenylalanine
Molecule contains single- and double-stranded regions
These spontaneously interact to produce this 3-D structure
Figure 9.24
