THREE: 1
UNIT GENETICS
C hapter
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
From Gene to Protein (Chapter Seventeen)
THE
THE CONNECTION BETWEEN GENES AND PROTEINS
The
The Study of Metabolic Defects Provided Evidence That Genes Specify Proteins
In
In 1909, Archibald Garrod suggested that genes dictated phenotypes
through enzymes that catalyze specific chemical reactions in the cell.
specific
How Genes Control Metabolism: One Gene-One Enzyme
Research
Research conducted several years later supported Garrods idea that a
gene dictates the production of a specific enzyme. Biochemists found
that cells synthesize and degrade most organic molecules via metabolic
pathways, in which each chemical reaction is catalyzed by a specific
enzyme.
A breakthrough in demonstrating the relationship between genes and enzymes occurred when Beadle
and Edward Tatum began working with a bread mold. After treating the mold with X-rays, they looked
ith
for mutants that differed from the wild-type in their nutritional needs. Wild-type bread mold can
type
survive
survive in the laboratory on agar mixed with inorganic salts, glucose, and biotin. They identified
mutants that needed certain essential molecules because they couldnt synthesize it themselves.
They found that their mutants fell into three classes, each mutated in a different gene. In the pathway
leading to synthesis of arginine, it was necessary to convert a precursor nutrient to orthinine, which is
ecessary
converted
converted to citrulline, which is converted to arginine. When they tested the mutants, their results
showed that the mutants were blocked at different steps in the pathway. Since each mutant had one
defective gene, their results provided strong support for the one gene-one enzyme hypothesis.
one
One Gene-One Polypeptide
Researchers
Researchers later learned that not all proteins are enzymes. Since proteins that are not enzymes are
still gene products, molecular biologists began to think in terms of one gene-one protein. However,
ecular
one
many
many proteins are made of two or more different polypeptide chains, with each polypeptide made by
its own gene. Beadle and Tatums idea can then be restated as the one gene-one polypeptide
o ne gene-one
hypothesis.
hypothesis
hypothesis
Transcription
Transcription and Translation Are the Two Main Processes Linking Gene to Protein: An Overview
Recall that RNA contains ribose instead of deoxyribose, has the base uracil instead of thymine, and is
usually single-stranded. Transcript ion is the synthesis of RNA under the direction of DNA. Just as a
Transcript
DNA
DNA strand provides a template for DNA synthesis of a complementary strand, it can provide a
template for assembling a sequence of RNA nucleotides. This type of RNA molecule is called
messenger
m ess
messenger RNA (mRNA), because it carries a message from the DNA.
mRNA),
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
Translation
Translation is the actual synthesis of a polypeptide, which occurs under the direction of mRNA.
During this stage, the cell translates the base sequence of mRNA into the amino acid sequence of a
polypeptide. Ribosomal RNA serves as the translation site.
In prokaryotes, DNA transcription and translation both take place in the cytosol. Thus, polypeptides
can be synthesized during translation. In a eukaryotic cell, the nuclear envelope separates
transcription from translation, which allows for RNA processing to yield the finished mRNA. An initial
RNA transcript is the primary transcript
transcript.
In the Genetic Code, Nucleotide Triplets Specify Amino Acids
Biologists established that triplets of nucleotide bases are the smallest units of
uniform length that can code for all 20 amino acids. If each arrangement of three
consecutive bases specifies an amino acid, there can be 64 possible code words.
Experiments have verified that there is a triplet code genetic instructions for a
code:
polypeptide gene are written in the DNA as a series of three-nucleotide words.
The intermediate step in translating a gene into amino acids is transcription, where
the gene determines the sequence of base triplets along the length of an mRNA
molecule. For each gene, the template strand of DNA is described (this strand is
also known as the antisense strand, because the RNA produced will be
complementary and exactly like the sense strand).
The mRNA base triplets are called codons Each codon stands for one of the 20 amino acids or it could
codons
ns.
be a stop codon that codes for a release factor. During translation, the sequence of codons are read in
the 5 3 direction along the mRNA.
Cracking the Genetic Code
The first codon was deciphered in 1961 by
Marshall Nirenberg, of the National Institutes of
Health. He synthesized an artificial mRNA of all
uracils to a test-tube mixture containing amino
acids and ribosomes. His artificial system then
created a polypeptide containing a single amnio
acid, phenalanine (Phe). All 64 codons were then
deciphered by the mid 1960s. The codon AUG is
the start codon, while also coding for methionine
(Met). UAA, UAG, and UGA are stop signals
that mark the end of translation.
While several codons can code for one amino acid, one codon never codes for more than one amino
acid. The correct reading frame is necessary to be able to create the right nucleotides as read from 5
3 in groups of three.
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
The Genetic Code Must Have Evolved Very Early In the History of Life
The genetic code is nearly universal. Genes can be transcribed and translated after being transplanted
from one species to another. Bacteria can be programmed by the insertion of human genes to
synthesize certain human proteins that have important medical uses. Exceptions to this universality
occur in certain single=celled eukaryotes, as well as certain mitochondria and chloroplasts. However,
the near universality indicates that the genetic code must have existed very early in the history of life.
THE SYNTHESIS AND PROCESSING OF RNA
Transcription is the DNA-Directed Synthesis of RNA: A Closer Look
mRNA, the carrier of information from DNA to the cells protein-synthesizing
machinery, is transcribed from the emplate strand of a gene. RNA
polymerase keeps the two strands of DNA apart and hooks together the
RNA nucleotides as the base-pair along the DNA template. Like the DNA
polymerases that function in DNA replication RNA polymerases can add
nucleotides only to the 3 end of the original strand.
Specific sequences of nucleotides along the DNA mark where transcription
of a gene begins and ends. The DNA sequence where RNA polymerase
attaches and initiates transcription is known as the promoter; the sequence
that signals the end of transcription is the terminator The stretch of DNA
terminator.
to be transcribed into an RNA molecules is called a transcription unit
unit.
RNA Polymerase Binding and Initiation of Transcription
The promoter of a gene includes within it the transcription start point and extends several dozen
nucleotide pairs upstream. The promoter also serves as the binding site for RNA polymerase and
determines which of the two strands of the DNA helix is used as the template.
In prokaryotes, the RNA polymerase itself specifically recognizes and binds to the promoter. In
eukaryotes, a collection of proteins called transcription factors mediate binding of RNA polymerase
and the initiation of transcription. RNA polymerase only binds to the promoter after certain
transcription factors are attached to it. The completed assembly of transcription factors and RNA
polymerase bound to the promoter is called a transcription initiation complex The TATA box is a
complex.
nucleotide sequence containing TATA (on the sense strand) that is about 25 nucleotides upstream from
the start point. A transcription factor that recognizes the TATA box will then bind to the DNA before
RNA polymerase does so. Additional transcription factors will join the polymerase on the DNA to form
the transcription initiation complex.
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
Elongation of the RNA Strand
As RNA polymerase moves along the DNA, it continues untwisting the
double helix to allow pairing with RNA nucleotides. The DNA double
helix re-forms and the new RNA molecule separates from the DNA
template.
A single gene can be transcribed simultaneously by several molecules of
RNA polymerase following each other. Cells can then make the encoded
protein in large amounts.
Termination of Transcription
Once the RNA polymerase reaches a terminator sequence in the DNA, transcription will end. In the
prokaryotic cell, transcription stops right at the end of the termination signal, releasing both the DNA
and RNA. In the eukaryotic cell, the polymerase continues for hundreds of nucleotides past the
termination signal and then cuts off.
Eukaryotic Cells Modify RNA after Transcription
Alteration of mRNA Ends
The 5 end of a pre-mRNA molecule is capped off with a modified form of a guanine nucleotide
(guanosine triphosphate). This 5 cap helps protect the mRNA from degration by hydrolytic enzymes in
the cytosol, and also functions as an attachment marker for ribosomes. The 3 end of mRNA receives a
poly(A) tail consisting of some 50 to 250
adenine nucleotides. This tail also inhibits
degradation of the RNA and probably helps
ribosomes to attach to it.
Split Genes and RNA Splicing
Large portions of the primary transcript are removed in RNA splicing Most eukaryotic genes and their
splicing.
RNA transcripts have long noncoding stretches of nucleotides that are interspersed between coding
segments of the gene. The noncoding segments are called intervening sequences or introns for short,
while the other regions are called exons
exons.
Researchers have discovered that signals for RNA splicing are short nucleotide sequences at the ends
of introns. Particles called small nuclear ribonucleoproteins, or snRNPs (pronounced snurps),
recognize these splice sites. snRNPs located are in the cell nucleus and are composed of RNA and
protein molecules. The RNA in a snRNP particle is called a small nuclear RNA (snRNA); each molecule
is about 150 nucleotides long. Several snRNPs join with additional proteins to form a spliceosome
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
that cuts at specific points to release introns, then immediately joins together the two exons once
adjacent to the intron. The idea of a catalytic role for snRNA arose from the discovery of ribozymes
ribozymes,
RNA molecules that function as enzymes.
Ribozymes
RNA is often involved in catalyzing reactions. In a few cases, splicing can occur completely without
proteins or even extra RNA molecules. For example, intron RNA in Tetrahymena splices itself.
The Functional and Evolutionary Importance of Introns
A number of genes are known to give rise to two or more different polypeptides, depending on which
segments are considered exons during RNA processing: this is called alternative RNA splicing Split
splicing.
genes may also facilitate the evolution of new and potentially useful proteins. Proteins often have a
modular architecture consisting of functional regions and structural regions called domains One
omains.
domain, for example, might include the active site, while another might attach the protein to a cellular
membrane. Different exons often code for different domains of a protein.
Introns increase the probability of potentially beneficial crossing over between genes. Introns increase
the possibility that a crossover could switch one version of an exon for another version found on the
homologous chromosome.
THE SYNTHESIS OF PROTEINS
Translation is the RNA-Directed Synthesis of a Polypeptide: A Closer Look
Transfer
tRNA),
Transfer RNA (tRNA transfers amino acids from the cytoplasm to a
tRNA
ribosome. Each molecule of tRNA carries a specific amino acid, as determined
by the anticodon which is part of the tRNA structure. The genetic message
anticodon,
will be translated as tRNA molecules deposite amino acids in the prescribed
order and the ribosome joins the amino acids into a chain.
The Structure and Function of Transfer RNA
A tRNA molecule consists of a single RNA strand about 80 nucleotides long.
There are about 45 types of tRNA, as some of them can recognize two or
more different codons. The relaxation of base pairing rules that allows for
multiple codons to code for the same amino acid is called wobble
wobble.
Aminoacyl-tRNA Synthetases
A tRNA that binds to an mRNA codon specifying a particular amino acid must cary only that acid to
the ribosome. Each amino acid joins to the correct tRNA by a specific enzyme called aminoacyl -tRNA
aminoacylsynthetase.
synthetase There are 20 of these enzymes in the cell, one enzyme for each amino acid. The
synthetase catalyzes the reaction between tRNA and an amino acid through the hydrolysis of ATP.
The resulting aminoacyl tRNA is called an activated amino acid.
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
Ribosomes
A ribosome is made up of two subunits, constructed
of proteins and RNA molecules named ribosomal
(rRNA).
RNA (rRNA) In eukaryotes, the sununits are
constructed inside the nucleolus. Once large and
small subunits attach to an mRNA molecule, they join
to form a functional ribosome.
Each ribosome has three binding sites for tRNA. The
P site holds the tRNA carrying the growing
polypeptide chain, while the A site holds the next
amino acid to be added to the chain. Discharged
tRNAs leave the ribosome from the E site.
Building a Polypeptide
Initiation.
I nitiation
Initiation The initiation stage brings together mRNA,
the first tRNA, and the two subunits of the ribosome.
First, a small ribosomal subunit binds to both mRNA
and the initiator tRNA, attaching to the leader
segment at the 5 end of the mRNA. The large
ribosomal subunit will then attach to the small
subunit. Proteins called initiation factors are required
to bring it all together. The cell also spends energy in
the form of a GTP molecule to form the initiation
complex.
Elongation.
Elongation Amino acids are added one by one in a chian. The mRNA codon in the A site forms
hydrogen bonds with the anticodon of an incoming tRNA (uses 2 molecules of GTP). The rRNA,
functioning as a ribozymes, will catalyze the formation of a peptide bond that joins the polypeptide
extending from the P site to the newly arrived amino acid. The ribosome now moves the tRNA in the A
site to the P site. The tRNA that was previously in the P site is moved into the E site, where it leaves
the ribosome. This move costs 1 GDP molecule.
Termination.
Termination Once a stop codon is reached, a protein called a release factorbinds directly to the stop
codon in the A site, which causes a water molecule to be added to the polypeptide chain. This
hydrolyzes the completed polypeptide from the tRNA in the P site.
Polyribosomes
Polyribosomes
Polyribosomes are strings of ribosomes that may trail along the same mRNA, which allow a cell to
make many copies of a polypeptide quickly.
From Polypeptide to Functional Protein
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
During and after synthesis, a polypeptide chain begins to coil and fold spontaneously, forming a
functional protein of specific conformation. Additional steps, posttranslational modifications, may be
required before the protein can begin functioning. Certain amino acids may be chemically modified,
while enzymes may remove one or more amino acids. A single polypeptide chain can also be cleaved
into two or more pieces.
Signal Peptides Target Some Eukaryotic Peptides To Specific Destinations In the Cell
Bound ribosomes are attached to the cytosolic side of rough endoplasmic reticulum. They work to
make proteins of the endomembrane system, as well as proteins secreted from the cell. Synthesis of
all proteins begins in the cytosol, when a free ribosomes starts to translate an mRNA molecule. The
process will continue unless the growing polypeptide cues the ribosome to attach to the ER. The
polypeptides of proteins destined for the endomembrane system or for secretion are marked by a signal
peptide,
peptide which targets the protein to the ER. The signal peptide will then be recognized as it emerges
from the ribosome by a signal- recognition particle (SRP This particle functions as an adapter that
ignalSRP).
SRP
brings the ribosome to a receptor protein built into the ER membrane.
Point Mutations Can Affect Protein Structure and Function
Mutations
Mutations are changes in the genetic material of a cell (or virus).
Point mutations are chemical changes in just one base pair of a
gene. If a point mutation occurs in a gamete, it may be
transmitted to offspring and to a succession of future
generations. If the mutation has an adverse effect on the
phenotype of a human or other animal, the mutant condition is
referred to as a genetic disorder, or hereditary disease.
The origins of sickle-cell disease can be traced to the mutation of
a single base pair in the gene that codes for one of the
polypeptides of hemoglobin.
Types of Point Mutations
Substitutions. baseSubstitutions
Substitutions A base- pair substitution is the replacement of one nucleotide and its partner in the
complementary DNA strand with another pair of nucleotides. Some substitutions are called silent
mutations because they have no effect on the encoded protein. Other changes of a single nucleotide
pair may switch an amino acid but have little effect on the protein. The new amino acid may have
properties similar to the one it replaced, or it may e in a region of the protein where the exact sequence
of amino acids is not essential to the proteins function.
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UNIT THREE: GENETICS
Chapter Seventeen: From Gene to Protein
(Text from Biology, 6th Edition, by Campbell and Reece)
However, base-pair substitutions in a crucial area such as the active site of an enzyme will significantly
alter protein activity. Occasionally, such a mutation will lead to an improved protein or one with novel
capabilities that enhance the success of the mutant organism and its descendants. But much more
often, these mutations are detrimental.
Substitution mutations are usually missense mutations
mutations;
that is, the altered codon will still code for an amino acid, but
not necessarily the right one. If a point mutation changes a
codon for an amino acid to a stop codon, it is known as
mutations.
nonsense mutations
Insertions and Deletions. Insertions and deletions are
additions or losses of nucleotide pairs in a gene. These
mutations have a disastrous effect because the insertion or
deletion may alter the triplet grouping of the genetic
mutation. Such a mutation, called a f ram eshif t m utat io n,
will occur whenever the number of nucleotides inserted or
deleted is not a multiple of three. All the nucleotides
downstream of the deletion/insertion will be improperly
grouped.
Mutagens
Errors during DNA replication, repair, or recombination can
lead to the mutations previously mentioned. Mutations
resulting from such errors are called spontaneous mutations.
A number of physical and chemical agents, called m utagen s, interact with DNA to cause mutations.
In the 1920s, Hermann Muller discovered that fruit flies subjected to X-rays had increased genetic
changes. Mutagenic radiation causes ultraviolet light that can produce disruptive thymine dimmers in
DNA. Chemical mutagens fall into several categories. Base analogues are chemicals that are similar to
normal DNA bases, but that pair incorrectly during DNA replication. Other chemical mutagens
interfere with correct DNA replication by inserting themselves into the DNA and distorting the double
helix. Other mutagens cause chemical changes to change their pairing properties.
Researchers have been able to develop various methods to test the mutagenic activity of different
chemicals. A major application is to test whether chemicals cause cancer. Since most carcinogens are
mutagenic and most mutagens are carcinogenic, this approach makes sense.