Trends in Biotechnology
Molecules of Genetics
Concept 15 - DNA and proteins are key
molecules of the cell nucleus.
DNA was discovered as a major chemical of the
nucleus at about the same time Mendel and
Darwin published their work. However, during
the early 1900s, more people thought that
proteins were the molecules able to carry large
amounts of hereditary information from
generation to generation.
Although DNA was known to be a very large
molecule, it seemed likely that its four chemical
components were assembled in a monotonous
pattern — like a synthetic polymer. Also, no
specific cellular function had yet been found for
DNA.
Proteins, on the other hand, were important as
enzymes and structural components of living
cells. Proteins were also known to be polymers
of numerous amino acids. These polymers are
called polypeptides. Most importantly, the 20
amino acid "alphabet" of proteins potentially
could be configured into more unique
information-carrying structures than the fourletter alphabet of DNA.
A nucleotide is
made of three
elements:
phosphate,
deoxyribose sugar,
and a nitrogenous
base.
The carbons of the
deoxyribose sugar are
numbered 1-5. In a
nucleotide, the
noitrogenous base is
always bound to
carbon#1, a hydroxyl
group (OH) is bound to
carbon#3 and the
phosphate group is
bound to carbon#5.
Each of the
four
nucleotides
has a
distinct
nitrogenous
base.
The sugars are
connected to the
phosphate group
through a
phosphodiester
bond.
The phosphodiester
bonds give the
molecule a
direction; from
carbon#5 to
carbon#3. [ 5 prime
(5’) to 3 prime (3’) ]
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Concept 16 - One gene makes one protein.
In 1902, Archibald Garrod described the
inherited disorder alkaptonuria as an "inborn
error of metabolism." He proposed that a gene
mutation causes a specific defect in the
biochemical pathway for eliminating liquid
wastes. The phenotype of the disease — dark
urine — is a reflection of this error.
Strong evidence for this hypothesis was found in
1941 by George Beadle and Edward Tatum, using
the simple bread mold Neurospora. First, they
found that molds exposed to radiation lose the
ability to produce essential nutrients, and this
slowed, even stopped the growth of the mold.
Then, they found that growth can be restored by
providing the mutated mold with a specific
supplement. They reasoned that each mutation
must inactivate the enzyme (protein) needed to
synthesize the nutrient. Thus, one gene carries the
directions for making one protein.
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Concept 17 - A gene is made of DNA.
In the 1920s, experiments showed that a
harmless strain of bacteria can become
infectious when mixed with a virulent strain of
bacteria that had been killed. The dead bacteria
apparently provide some chemical that
"transforms" the harmless bacteria to infectious
ones. This so-called "transforming principle"
appeared to be a gene.
A team of scientists led by Oswald Avery,
followed up on these experiments in the 1940's.
They found that a pure extract of the
"transforming principle" was unaffected by
treatment with protein-digesting enzymes but
was destroyed by a DNA-digesting enzyme. This
showed that the transforming principle is DNA
— and, by extension, a gene is made of DNA.
Still, many scientists were slow to accept this
clear proof that DNA, not protein, is the genetic
molecule.
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Concept 18 - Bacteria and viruses have DNA too.
Microscopes proved the existence of single-celled
bacteria. However, there was debate about
whether bacteria had genes and what attributes
they may have in common with higher life forms.
This debate was settled in the 1940's, when it was
discovered that bacteria have sex. During the
process of conjugation, genes are exchanged
through a mating channel that links two bacteria.
Electron microscopy suggested that bacterial
viruses carry on a similar process. A virus
attaches to a host bacterium and injects its
genes through its channel-like tail. In 1952,
Alfred Hershey showed that DNA, alone, is
responsible for the reproduction of new viruses
within an infected cell. This provided undeniable
support for Avery's earlier experiments that a
gene is made of DNA. It also showed that
viruses, as well as bacteria, can be used as
models for studying universal principles of
genetics.
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Concept 19 - The DNA molecule is shaped like a
twisted ladder.
Earlier work had shown that DNA is composed of
building blocks called nucleotides consisting of a
deoxyribose sugar, a phosphate group, and one of
four nitrogen bases — adenine (A), thymine (T),
guanine (G), and cytosine (C). Phosphates and
sugars of adjacent nucleotides link to form a long
polymer. Other key experiments showed that the
ratios of A-to-T and G-to-C are constant in all living
things. X-ray crystallography provided the final clue
that the DNA molecule is a double helix, shaped like
a twisted ladder.
In 1953, the race to determine how these pieces
fit together in a three-dimensional structure was
won by James Watson and Francis Crick at the
Cavendish Laboratory in Cambridge, England.
They showed that alternating deoxyribose and
phosphate molecules form the twisted uprights
of the DNA ladder. The rungs of the ladder are
formed by complementary pairs of nitrogen
bases — A always paired with T and G always
paired with C.
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Concept 20 - A half DNA ladder is a template for
copying the whole.
Because of the pairing of adenine-to-thymine and
guanine-to-cytosine, Watson and Crick proposed
that one half of the DNA ladder can be a template
for recreating the other half during DNA replication.
By 1958, two lines of evidence came together to
provide proof of this hypothesis. First, an enzyme
was discovered — DNA polymerase — that adds
complementary nucleotides to the template
provided by a half DNA molecule.
Second, an ingenious experiment used nitrogen
isotopes to follow the construction of new DNA
molecules during successive generations of
bacteria. This showed that one strand of each
DNA molecule is passed along unchanged to
each of two daughter cells. This "conserved"
strand acts as the template for DNA polymerase
to synthesize a second complementary strand,
which completes each new DNA molecule.
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Concept 21- RNA is an intermediary between DNA
and protein.
DNA is found mostly in the cell nucleus, but another
type of nucleic acid, RNA, is common in the
cytoplasm. Watson and Crick proposed that RNA
must copy the DNA message in the nucleus and
carry it out to the cytoplasm, where proteins are
synthesized. Crick also predicted the existence of an
"adaptor" molecule that reads the genetic code and
selects the appropriate amino acids to add to a
growing polypeptide chain. This proposed flow of
genetic information from DNA to RNA to protein
became known as the "Central Dogma."
As it turned out, several types of RNA are
involved in the utilization of genetic information.
In the nucleus, the DNA code is "transcribed," or
copied, into a messenger RNA (mRNA) molecule.
In the cytoplasm, the mRNA code is "translated"
into amino acids. Translation is orchestrated at
the ribosome — itself partly composed of RNA
— with transfer RNA playing the role of adaptor.
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Information flows between DNA, RNA and
protein. DNA -> protein is another special
transfer, but it is not found in nature.
Concept 22 - DNA words are three letters long.
The genetic code had to be a "language" —
using the DNA alphabet of A, T, C, and G — that
produced enough DNA "words" for each of the
20 known amino acids. Only 16 words are
possible from a two-letter combination, but a
three-letter code produces 64 words.
Researchers assumed a three-letter code called
a codon.
Research teams carefully synthesized different
RNA molecules, each a long strand of a single
repeated codon. Then, each type of synthetic
RNA was added to a cell-free translation system
containing ribosomes, transfer RNAs, and amino
acids. Each type of synthetic RNA produced a
polypeptide chain of repeated units of a single
amino acid. Several codons are "stop" signals
and many amino acids are specified by several
different codons. All 64 three-letter
combinations do something.
In 1961, Marshall Nirenberg and J. H. Matthaei
published their paper. They showed that a
synthetic messenger RNA made of only uracils
can direct protein synthesis. The polyU mRNA
resulted in a poly-phenylalanine protein They
had the first piece of the genetic code.
Later, Nirenberg and his group found the entire
genetic code by matching amino acids to
synthetic triplet nucleotides. There is
redundancy (some amino acids are encoded by
more than one codon) and some codons are
"punctuation marks" in the mRNA message.
They also showed that with few exceptions, the
genetic code was the same for all organisms.
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Concept 23 - A gene is a discrete sequence of DNA
nucleotides.
Mendel described a gene as a discrete unit of
heredity that influences a visible trait. Beadle and
Tatum defined a gene as the discrete directions for
making a single protein, which influences a
metabolic trait. Early sequencing efforts showed
that proteins are, in turn, long chains of amino acids
arranged in a specific order. The triplet genetic code
further refined the definition of a gene as a discrete
sequence of DNA encoding a protein — beginning
with a "start" codon and ending with a "stop"
codon.
Gene analysis took a giant step forward with the
discovery of methods to determine the exact
sequence of nucleotides that compose a specific
gene. DNA sequencing was built upon earlier
knowledge of DNA polymerases and cell-free
systems for replicating DNA. The chaintermination method, which makes clever use of
a "defective" DNA nucleotide, now dominates
DNA sequencing technology.
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Concept 24 - The RNA message is sometimes
edited.
Dogma and logic dictated that the mRNA code is
a faithful representation of the DNA from which
it is transcribed. This exact correspondence
between mRNA sequence and DNA sequence
was generally upheld in experiments with
bacterial cells (prokaryotes).
However, inconsistencies surfaced as
recombinant-DNA techniques allowed
researchers to explore the genes of higher cells
(eukaryotes). Then, it was found that mRNA
transcripts appeared to be shorter than their
corresponding genes. This difference became
obvious in electron micrographs of mRNA bound
to its complementary DNA template — where
regions of DNA without corresponding mRNA
form loops.
In fact, the protein coding information in genes
is interrupted by non-coding sequences called
introns, which results in "split genes." The entire
DNA code is faithfully transcribed into a
temporary form of RNA (pre-mRNA), but this is
edited in the nucleus to yield a mature mRNA.
The process of RNA splicing involves removing
non-coding regions, introns, and splicing
together adjacent coding regions, exons.
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Concept 25 - Some viruses store genetic
information in RNA.
DNA was believed to be the sole medium for
genetic information storage. Furthermore,
Watson and Crick's central dogma assumed that
information flowed "one-way" from DNA to RNA
to protein. So it came as a surprise when in
1971, it was discovered that some viruses shift
their genetic information from RNA to DNA.
Even so, these viruses ultimately make proteins in
the same way as higher organisms. During infection,
the RNA code is first transcribed "back" to DNA —
then to RNA to protein, according to the accepted
scheme. The initial conversion of RNA to DNA —
going in reverse of the central dogma — is called
reverse transcription, and viruses that use this
mechanism are classified as retroviruses. A
specialized polymerase, reverse transcriptase, uses
the RNA as a template to synthesize
complementary and double-stranded DNA
molecule.
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Concept 26 - RNA was the first genetic
molecule.
Experiments in the 1960s showed that
messenger RNA has the ability to store genetic
information, while transfer and ribosomal RNA
have the ability to translate genetic information
into proteins. Experiments performed two
decades later showed that some RNAs can even
act as an enzyme to self-edit their own genetic
code!
These results raised
two questions: 1) Why
does RNA play so many
roles in the flow of
genetic information?
2) Why bother storing
genetic information in
DNA, if RNA alone
could do the job?
RNA has great capability as a genetic molecule;
it once had to carry on hereditary processes on
its own. It now seems certain that RNA was the
first molecule of heredity, so it evolved all the
essential methods for storing and expressing
genetic information before DNA came onto the
scene. However, single-stranded RNA is rather
unstable and is easily damaged by enzymes. By
essentially doubling the existing RNA molecule,
and using deoxyribose sugar instead of ribose,
DNA evolved as a much more stable form to
pass genetic information with accuracy.
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