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CHAPTER
22
THE CHEMICAL NATURE OF THE GENE
We have seen how studies of inheritance led to the idea
that the characteristics of an organism are controlled by
individual elementscalledgenes.We'vealso explored some
aspectsof biological chemistry and examined the kinds
of molecules found in living cells. Is it possible that genes
are made of ordinary molecules?Could a molecule actually
carry genetic information? In this chapter we will try to
answer these questions.
CodingCapacity:
ProteinsSeemto Bethe BestBet
Could protein be the genetic material? Eukaryotic chromosomescontain very little carbohydrate; they are about
30 percent nucleic acid and from 60 to 75 percentprotein.
Most scientists in the first half of the twentieth century
agreed that proteins were most likely to be the molecules
that carry information. To begin with, protein molecules
are composed of 20 amino acids, but nucleic acids are
composedof just 4 nucleotides.Therefore, proteins should
be better coding molecules (an alphabet with 20 lette~s
~ould be far more exgressive than one with just.::JJ.Proteins also differ a great deal more from one species to
another than nucleic acids do. Finally, although scientists
at the time were familiar with the ability of protein enzymes
to control and regulate chemical reactions, there was no
evidence that nucleic acids were able to do anything.
The Transforming Principle
As early as the 1920s, a few people were working on
projects that were eventually to identify the molecules
responsible for inheritance. Frederick Griffith wasa British scientist studying the way in which certain types of
408
livesmooth
lethal
Figure 22.1 A diagrammatic summary of Griffith's
experiments on transformation. Something present
in a heat-killed extract of virulent pneumonia bacteria was able to transform a nonvirulent strain of
bacteria so that they caused pneumonia. Griffith
recovered live, virulent bacteria from the infected
mice.
injection
,~~
8!P;8
live rough
harmless
injection
heat-killed
smooth
harmless
injeLiion
fIf~8
(psI
!kft~z:~~
blood from dead mouse
~
combined heat-killed
smooth and live rough
bacteria causepneumonia,a serious and often fatal lung
disease.
In 1928 Griffith had in his laboratory two different
types-strains-of pneumonia bacteria. Both strains grew
very well in special culture plates in his lab, but only one
of them actually caused pneumonia in mice. Griffith
managed to grow both strains succ~ssfullyin the lab and
noticed that he could distinguish one strain from the
other simply by its appearance on a culture dish. The
virulent (disease-causing)bacteria produced a jelly-like
coating around themselvesthat made their colonies look
smooth. The nonvirulent (harmless)strainsmade no such
coating and instead grew into colonies with roughO'jagged edges.
Griffith first tested to determine whether the coating
might contain a disease-causingpoison. To do this, he
killed a culture of virulent cells and injected the dead
cells (coatings and all) into the mice. The mice were not
harmed by the injections. Griffith concluded that his suspicions about a poison were incorrect. Next he injected
both live nonvirulent and killed virulent cellsinto a mouse
(neither one of these injections by itself made any of the
mice sick). To Griffith's surprise, when the two types of
cells were injected at the same time, the mice developed
pneumonia (Figure 22.1).
Today it is hard to appreciate just how startling this
result was. The live nonvirulent bacteria never made the
mice sick, and neither did the killed virulent ones. Why
should the combinationof these two have an effect that
was different from what was seen when the two were
administered separately?
To confuse matters further, Griffith recovered live
bacteria from the animals that had developed the disease.
Were these bacteria the same nonvirulent ones he had
injected? He grew the bacteria on plates to find out. Now
they formed the smooth colonies that were characteristic
of the virulent strain. Griffith:Sextracthad transformedone
kind of bacteriuminto another! Griffith called the process
he had discovered transformation.
Griffith spe~ulated that when the live and killed bacteria were mixed together, a "factor" was transferred from
the killed cells into the live ones. This factor changed the
characteristicsof the live cells in a permanent way,so that
they henceforth acted like the virulent ones. What had
actually happened? The moleculeof inheritancehad been
transferred,and it had changedthe characteristics
of the cell
that receivedit.
The Transforming Principle is DNA
In 1942 Oswald Avery, a scientist at the Rockefeller Institute in New York City, devised a simple research project:
He would determine the chemical nature of the material
CHAPTER
22
Molecules and Genes
409
Figure 22.2 Thebasicstructure of a nucleic acid is illustrated in this dinucleotide. Both
RNA and DNA consistof long
chains in which the sugar and
phosphate groups of adjacent
nucleotidesare joined by covalent bonds.
responsible for the transformation effect in Griffith's
experiments. Using Griffith's transformation system,
Avery and his co-workerstreated the transforming extract
in waysthat destroyed proteins, carbohydrates,lipids, and
a kind of nucleic acid known as ribonucleic acid (RNA).
Transformation still occurred. However,when they treated
the transforming extract with an enzyme that destroyed
another type of nucleic acid, deoxyribonucleicacid (DNA),
transformation did not occur. Proteins did not carry
genetic information. DNA did.
Avery's results were treated with som,s!epticism)by
his colleagues,if only becausehis answerseemedto make
so little sense.Many in the scientific community had not
suspectedthat nucleic acids were capable of playing such
an important role.
Erwin Chargaff, a biochemist, was the first to notice a
pattern in the relative percentagesof the four bases.This
pattern is apparent in Table 22.1.
.
Do you seethe same p~ttern that Chargaff did? The
percentages of the four basesare not static. They vary
over a wide range, but the relative proportions of guanine (G) and cytosine (C) are always nearly equal; the
sameis true for the proportions of adenine (A) and thymine (T). In more symbolic form, we might express this
observation as follows:
[A]
[C]
= [T]
= [G]
This observation became known as Chargaff's rule.
Chargaff himself had no explanation for the rule, but it
did suggest something about the structure of DNA.
Becausethere were alwaysequal amounts of adenine and
thymine, for example, the molecule had to be organized
in a way that would account for the equivalence of the
two bases. Therefore, to our basic knowledge of the
chemical structure of a polynucleotide we can add a bit
of information about the ratios of the various bases.
The X-ray Pattern
If a molecule can be crystallized, X-ray diffraction may
produce a scattering pattern from the crystal that yields
information about the molecule's internal structure. The
repeating pattern of atomic bonds that exists within a
crystal scatters a beam of X rays in a regular pattern.
When researchers realized that DNA might be an interesting and important molecule, a number of X-ray crystallographers began to work on its structure. In most
cases,however, the formation of good crystals from DNA
turned out to be both a practical and a theoretical problem. Crystals were difficult to produce, and the patterns
from successfulcrystals were difficult to interpret.
CLUESTO THE STRUCTURE OF DNA
BaseComposition
We have already seen that nucleic acids are polymers of
nucleotides. Long chains, polynucleotides, can be built
by linking nucleotides together (Figure 22.2). By the late
1940s,scientistsunderstood the general chemistry of the
polynucleotide. Still there was nothing that might distinguish the nucleic acid from any other biological polymer.
Some of the first clues to the structure of DNA came
from experiments in which scientists determined the
compositionof DNA obtained from different organisms.
DNA is a nucleic acid containing four nucleotide bases:
adenine, cytosine, guanine, and thymine. Each of these
basesis represented by a single letter: A, C, G, and T.
410
PART
5
Molecules of Ufe
Table 22.1 BaseComposition (% of Total DNA Bases)
SOURCE:
Academic
E. Chargaff
Press.
and J. Davidson,
eds., 1955, The Nucleic Acids, New York:
A few scientists,however, believed that DNA wassuch
an interesting molecule that X-ray diffraction should be
attempted even if perfect crystals couldn't be formed.
One such person was Rosalind Franklin, a young scientist
working with Maurice Wilkins, a crystallographer in London. Franklin drew a thick suspension of the fiber-like
DNA molecules up into a glass capillary tube and used
this DNA sample to scatter X rays. In the tube, she hoped,
the thick suspension of DNA molecules would be forced
to line up so that the molecules were parallel to the tube.
Like spaghetti drawn through a straw,the moleculeswere
all arranged in the same direction-not perfect enough
to give a crystal-like pattern, but just good enough to
yield a few cluesabout the structure of the DNA molecule.
Interpreting
the X-rayPattern
One of the X-ray patterns produced by Franklin's DNA
samples is shown in Figure 22.3. The pattern contains
two critical clues to the structure of the DNA molecule
(graphically summarized 'in the figure).
Clue number1: The two large dark patches at the top
and bottom of the figure showed that some structure in
the molecule was arranged at a right angle to the long
axis of DNA and repeated at a distance of 3.4
A (see Ap-
pendix: The Metric System). In other words, something
in the molecule was arranged l~kethe run){s of a ladder.
Clue number2: The X-like mark in the center of tlie
pattern showed that something in the molecule was
Figure 22.3 (left) DNA fibers were taken up in a thin
tube so that most of them were oriented in the same
direction. An X-ray diffraction pattern was then
recorded on film. (bottom) X-ray diffraction pattern of
DNA in the "8" form, as taken by Rosalind Franklin in
1952. Franklin's X-ray pattern contained two important
clues to the structure of DNA. The large spots on the
top and bottom of the pattern indicate that there is a
regular spacing of 3.4 A along the length of the fiber.
The "X"-shaped pattern in the center indicates that
there is a zigzag feature in the molecule, which might
be consistent with a helix.
C HAP
T E R
22
Molecules and Genes
411
which dominated the picture could arise only ftom a helical structure."
Watson and Crick had to account for several things.
An ideal model for the structure of DNA would:
I.
2.
3.
4.
5.
Explain Chaigaff's rule.
Be able to carry coded information.
Be capable of replication.
Fit the chemistry known for polynucleotides.
Agree with the X-ray pattern's three predictions:
(a) The molecule is helical.
(b) One feature of the molecule is stackedat a spacing
of 3.4 A.
(c) The width of the molecule is abOut 20 A.
Becausethe X-ray data were not detailed enough to
determine a structure directly, Watson and Crick hoped
to find an arrangement of nucleotide subunits that would
be consistent with each piece of the puzzle, including the
X-ray pattern.
The Double Helix Model
Figure 22.4 There were many false steps along the road to
solving Franklin's X-ray pattern. For example, placing the nitrogenous bases inside the sugar-phosphate chains would give a
fiber of the correct average width, but the different sizes of the
bases, if there were no restrictions on how they were placed,
would produce a "Iumpy" fiber.
arranged in a zigzag fashion at a spacing of about 20 A.
If DNA had a twisting, helical configuration, the X-ray
pattern it produced from the side would indeed look like
that "X." The X-ray pattern suggestedthat the molecule
was a helix and that its diameter was about 20
A.
The scientific challenge remaining was to put all of
these clues together to determine the structure of DNA.
In 1953 two young scientistsworking at Cambridge University in England learned about Franklin's remarkable
X-ray pattern. The scientists,James Watson and Francis
Crick, had been working with molecular models, twisting
little bits of wood and paper into various shapes in an
effort to determine how nucleotides could form a structure that could do all that DNA wasable to do. According
to their own accounts of that discovery, one look at
Franklin's X-ray pattern was the last bit of evidence they
required, and the problem was solved.
In his book The Double Helix, James Watson wrote,
"The instant I saw the picture my mouth fell open and
my pulse began to race. . . . The black crossof reflections
412
PART
5
Molecules of Life
In early 1953 Watson and Crick believed that they had a
sensible structure for the molecule: the double helix
model. They published their ideas in a brief paper that
appeared in April of that year.The details of their model
were surprisingly simple. Watson and Crick realized they
could account for the 3.4 A spacingin the X-ray pattern
if they arranged the nitrogenous basesso that they were
"stacked" on top of each other.
The 20 A width of the molecule could be accounted
for if they placed two antiparallel strands side by side in
opposite directions and arranged the bases facing each
other.
The helical twist that was evident in the diffraction
patterns could be accounted for as well. AU they had to
do was twist the molecule so that the two strands twisted
about each other.
At first, however, there were two problems with the
model. First, what kinds of forces might hold the two
strands together? Second, how could one solve the problems posed by the sizesof the nitrogenous bases?Two of
the bases, adenine and guanine, belong to a chemical
group known as the purines. They have two carbon-nitrogen rings in their basic structures. The other
two, thymine and cytosine, are pyrimidines: They have
a single ring, meaning that they are quite a bit smaller
than the purines. This would cause a problem in the
model. If two pyrimidines were paired, the two strands
would have to be much closer than when two purines
were paired, making the model "lumpy" (Figure 22.4).
Chargaff's rule showed how a "lumpy" helix could be
avoided. If a purine was alwayspaired with a pyrimidine,
the helix wouldn't be lumpy. But wasit possible for bonds
to form between purines and pyrimidines to hold the two
strands together? Watson and Crick remembered how
hydrogen bonds (weak, noncovalent interactions) seemed
to stabilize the structure of the a helix in proteins. To
their delight, when James Watson drew a sketch of the
bases,they could find perfect placesfor hydrogen bonds
to form between A and T and between G and C (Figure
22.5).
The specific hydrogen bonding between the basesis
known as base pairing. ~atso!!,by the way, had nev~r
done very well in ch!!:!!!:-istry.
and his sketch~owed~e
ri~~onc~hird
hxdrogen bond (highlighted in
Figure 22.5) existed between G and C. Watsonand Crick's
insight solved one of the critical problems regarding the
biolOgicalrole of DNA. Prior to 1953, no one had been
able to come up with a reasonable scheme for how a
molecule might be replicated.But the structure of DNA
itself contained an obvious answer to the riddle. Each
strand is a complementary"copy" (although not an exact
copy) of the other, which means that each contains the
"information" required to reproduce the other strand.
All that is required to copy the molecule is to separate
the two strands and then to form a new strand for each
original one by using the base-pairing rules suggestedby
Watson and Crick. James Watson and Francis Crick (Figure 22.6) had put together a three-dimensional model
that accounts for the biological properties of DNA (Figure 22.7; see Theory in Action, The Prize, p. 414.)
H
/
N\
0
0 ..
H
..
H '\
H
H/' -H
"H
Figure 22.5 The problem of placing nitrogenous bases was
solved by James Watson, who drew sketches (top) showing how
hydrogen bonds might pair thymine with adenine and guanine
with cytosine. The modern representation of A- T and G-C base
pairing (bottom) shows three hydrogen bonds in the G-C pair,
something that was missing from Watson's first sketch.
Figure 22.6 (left) JamesWatsonand Francis
Crick shortlyafterthe publicationof their
double helix model for the structure of DNA.
(right) The two strandsof a DNA double helix
are held together by hydrogen bonds
between the nitrogenous bases.The strands
run in antiparallel directions, forming a helix
with a diameter of 20 A.
C HAP
T E R
22
Molecules and Genes
413
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THEORY
N
ACT
0 N
ThePrize
TheX.ray pattern that provided the
critical c1uesJorWatson and Crick
was made by Rosalind Franklin, Her
papeiw hichcQn tainedthepa ttern.
was actually published in the very
same issueofNatUfethat..<;ontained
the double heliX paper, Needlessto
s~y.she was norexactlyovedQycd that
a detai ledinte rp tetationo(hetown
tarieously
data was, publishedsimul
c
c
c
c
c
wit\) theda~itself,
c;
"c
WatsonandCnc
CCC
k
c
C
C
C
C
CCCC
havenev~rc
_
CCC
deniedthat Fra~kIin'spatterncohtairiedtheim~ttant newinfQrmation.
,;
that madethe.1rcmodel
possible,But
C
..
'.
C
their acuons,Ihterpreu~gthedata
aftetanadvan~c
" look.wet~~thical,c
b'
h
h C
0 ne 0~ f Fran.kl inS
)ogr~p ers as
414
PAR
T
Molecule~ of Life
argued that Watson and Crick
deprived Franklin of the credit she
deserved for making one of the fundamental scientific discoveries of the
twentieth century.
Unfortunately, Rosalind Franklin
lived long enough to see only the first
hint of how important her work had
been. She died of cancer in 1957,just
as DNA was beginning to become a
central focus in biological research. A
few years later, the Nobel Prize committee decided that the time was right
to award the Prize to the group that
had developed the double helix
model. The Nobe.lPrize is never
givepposthumously. It was awarded
to Watson and Crick, and Franklin's
associate, Maurice Wilkins.
~
DNA REPLICATION
The DNA molecule, as we see, suggestsa method for its
own replication. Watson and Crick pointed this out in a
paper published shortly after the announcement of the
double helix structure.
Figure 22.8 illustrateshow the double helix is unwound
to enable each strand to serve as the templatefor the synthesis of a new strand. The rules of complementary base
pairing help to control the proeessand ensure that each
newly synthesized strand has the appropriate base
sequence.
The replication process is said to be semiconservarive, which means that the two original strands of the
helix are separated and that at the end of replication,
each strand is paired with one of the newly synthesized
strands. In this way,the replication processproduces two
identical DNA molecules, each composed of one "old"
strand and one "new" strand.
5'
3
---8Parentmolecule
;8
Original strands
are unwound
DNA
polymerase
complex
"'\
3
y
5'
of
direction
DNA Polymerase
\
.
It was soon discovered that DNA does not replicate in
isolation but rather requires a number of specialenzymes
to unwind the double helix and synthesize new DNA
strands. The most important enzyme in this process is
known as DNA polymerase. The action of DNA polymerase is illustrated in Figure 22.8. DNA polymerase
contains a binding site for attachment to the DNA strand
and also a binding site for nucleotides. The nucleotides
that attach to the enzyme are triphosphates:
Like ATP, they
have three phosphate groups attached to them. DNA
polymerase binds the correct nucleotide to the growing
DNA strand and uses the energy from splitting off two
of the three phosphates to form a covalent bond linking
the nucleotide to the growing chain. As DNA polymerase
moves along the chain to attach the next molecule, part
of the enzyme "proofreads" the work it hasjust done by
checking the nucleotide pair to ensure that the proper
base pairing has taken place. If an incorrect nucleotide
has been inserted, this portion of the enzyme swiftly
removes the nucleotide from the chain, and the molecule
starts work over again.
This intricate process of base pairing, covalent bond
formation, and proofreading is at the heart of DNA replication in all organisms. In tiny bacteria, a few DNA
polymerase molecules may work to replicate the single
DNA molecule that contains all of the cell's genetic information. In larger organisms, thousands of DNA polymerasemolecules may work at scattered sitesthroughout
many chromosomes to complete DNA replication in time
for cell division to begin.
synthesis
"
e Complementary
'.,.
newstrands-synthesized
usingbase
sequence
of
originalstrands
3'
5'
DNA
polymerase
3'
-~
'/
-
~
complex
Figure 2~.8 Because
each strand of the DNA molecule is complementary to the other strand, each may serve as a template
against which a new strand may be constructed. DNA replicates
in semi-conservative fashion. Each strand of the helix serves as a
template against which a new strand is assembled, following
base-pairing rules.
C HAP
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Molecules and Genes
415