Lecture 2

Molecular Structure of DNA and RNA part 2

Chapter 9, pages 237 - 250

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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

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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

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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

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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

Rosalind Franklin

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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

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Testing the Hypothesis

Refer to Figure 9.14

9-37Figure 9.14

9-38Figure 9.14

The Data

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Interpreting the Data

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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

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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

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Figure 9.15

This hypothesis was, of course, incorrect!

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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

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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 Rosalind Franklin died in 1958, and Nobel prizes are not

awarded posthumously

Watson and Crick

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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

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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

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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

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Figure 9.17

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Figure 9.18

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The DNA double helix can form different types of secondary structure

The predominant form found in living cells is B-DNA

However, under certain in vitro conditions, A-DNA and Z-DNA double helices can form

DNA Can Form Alternative Types of Double Helices

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A-DNA Right-handed helix 11 bp per turn Occurs under conditions of low humidity Little evidence to suggest that it is biologically important

Z-DNA Left-handed helix 12 bp per turn Its formation is favored by

GG-rich sequences, at high salt concentrations Cytosine methylation, at low salt concentrations

Evidence from yeast suggests that it may play a role in transcription and recombination

9-52Figure 9.19

Bases substantially tilted relative to the central

axis

Bases substantially tilted relative to the central

axis

Sugar-phosphate backbone follows a

zigzag pattern

Bases relatively perpendicular to the

central axis

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In the late 1950s, Alexander Rich et al discovered triplex DNA It was formed in vitro using DNA pieces that were made

synthetically

In the 1980s, it was discovered that natural double- stranded DNA can join with a synthetic strand of DNA to form triplex DNA The synthetic strand binds to the major groove of the

naturally-occurring double-stranded DNA Refer to Figure 9.20

DNA Can Form a Triple Helix

9-54Figure 9.20

Triplex DNA formation is sequence specific

The pairing rules are

Triplex DNA has been implicated in several cellular processes

Replication, transcription, recombination

Cellular proteins that specifically recognize triplex DNA have been recently discovered

T binds to an AT pair in

biological DNA

C binds to a CG pair in

biological DNA

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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

This topic will be discussed in detail in Chapter 10

The Three-Dimensional Structure of DNA

9-56Figure 9.21

DNA wound around histone

proteins

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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

9-58Figure 9.22

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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 the A form with 11 to 12 base pairs per turn

Different types of RNA secondary structures are possible Refer to Figure 9.23

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Figure 9.23

Also called hair-pin

Complementary regions

Noncomplementary regions

Held together by hydrogen bonds

Have bases projecting away from double stranded regions

9-61

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

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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