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AP Biology Ch. 17
From Gene to Protein
Inherited instructions in DNA
direct protein synthesis.
The study of metabolic defects provided
evidence that genes specify proteins.
-Archibald Garrod was the first to propose
the relationship between genes and proteins
(1909). He suggested that genes dictate
phenotypes through enzymes that catalyze
reactions. Inherited diseases (inborn errors
in metabolism) reflect the person’s inability
to make particular enzymes.
How Genes Control Metabolism
• Geneticists George Beadle and Boris
Ephrussi (1930s) were able to demonstrate
the relationship between genes and enzymes
by studying eye color in Drosophila and
Beadle and Edward Tatum studied
mutations of a bread mold, Neurospora.
• They found that mutants of the bread mold
could not survive on minimal medium
because they lacked the ability to synthesize
essential molecules.
Terms:
• Auxotroph=nutritional mutants that can
only be grown on minimal medium
augmented with nutrients not required by
the wild type.
• Minimal medium=Support medium that is
mixed only with molecules required for the
growth of wild-type organisms.
• Complete growth medium=Minimal
medium supplemented with all 20 amino
acids and some other nutrients.
Beadle and Tatum: Conclusions
• Beadle and Tatum identified specific metabolic
defects from mutations by transferring fragments of
auxotrophic mutants growing on complete growth
medium to vials containing minimal medium each
supplemented with only 1 additional nutrient.
• For example: if a mutant grew on minimal medium
supplemented with only arginine, it could be
concluded that the mutant was defective in the
arginine synthesis pathway.
One Gene--One Polypeptide
• Beadle and Tatum’s one gene-one enzyme
hypothesis has been slightly modified:
– while most enzymes are proteins, many
proteins are not enzymes but are still gene
products.
– Many proteins are composed of 2 or more
polypeptide chains, each chain specified by a
different gene.
Transcription--Translation
• Transcription and translation are the 2 main
steps from gene to protein:
– RNA links DNA’s genetic instructions for
making proteins to the process of protein
synthesis. It copies or transcribes the message
from DNA and then translates that message into
a protein.
– The linear sequence of nucleotides in DNA
ultimately determines the linear sequence of
amino acids in a protein.
DNA/RNA Comparison
• DNA
• Double stranded
molecule of nucleic
acid
• Deoxyribose is the 5carbon sugar.
• Consists of 4 nitrogen
bases: adenine,
guanine, cytosine, and
thymine.
• RNA
• Single stranded
molecule of nucleic
• Ribose is the 5-carbon
sugar
• Consists of 4 nitrogen
bases: adenine,
guanine, cytosine, and
uracil
Transcription--Translation
• Transcription=The synthesis
of RNA using DNA as a
template
– A gene’s unique
nucleotide sequence is
transcribed from DNA to
a complementary
nucleotide sequence in
messenger RNA (mRNA).
– The resulting mRNA
carries this transcript of
protein-building
instructions to the
ribosomes.
• Translation=Synthesis of a
polypeptide, which occurs
under the direction of mRNA
– The linear sequence of
bases in mRNA is
translated into the lienar
sequence of amino acids in
a polypeptide.
– Translation occurs on
ribosomes, complex
particles composed of
ribosomal RNA (rRNA)
and protein.
Prokaryotes vs Eukaryotes
• Prokaryotes lack
nuclei, so DNA is not
segregated from
ribosomes or the
protein synthesizing
machinery. Thus,
transcription and
translation occur in
rapid succession.
• Eukaryotes have
nuclear envelopes that
segregate transcription
in the nucleus from the
translation in the
cytoplasm; mRNA is
modified before it
moves from the
nucleus to the
cytoplasm where
translation occurs.
The Genetic Code
• In the genetic code, a particular triplet of
nucleotides specifies a certain amino acid.
• Researchers have verified that the flow of
information from a gene to a protein is
based on a triplet code.
• Triplets of nucleotides are the smallest units
of uniform length to allow translation into
all 20 amino acids with plenty to spare.
• There 3-nucleotide “words” are called
codons.
Codons
• Codon=A 3-nucleotide sequence in mRNA
that specifies which amino acid will be
added to a growing polypeptide or that
signals termination; the basic unit of the
genetic code.
• Genes are not directly translated into amino
acids, but are first transcribed as codons in
mRNA.
• For each gene, only one of the 2 DNA
strands is transcribed (the template strand).
An mRNA is complementary to the DNA
template from which it is transcribed.
• If the triplet nucleotide sequence on the
template DNA strand is CCG; GGC, the
codon for glycine, will be the
complementary mRNA transcript.
• Remember: uracil (U) in RNA is used in
place of thymine (T) of DNA.
• Each mRNA codon specifies which one of
20 amino acids will be incorporated into the
polypeptide.
Cracking the Genetic Code
• 1961--Marshall Nirenberg of the National
Institute of Health deciphered the first
codon.
• All 64 codons have since been determined.
– 61 of the 64 codons code for amino acids.
– The triplet AUG has a dual function--it is the
start signal for translation and codes for
methionine.
– 3 codons do not code for amino acids, but
signal termination (UAA, UAG, and UGA).
Redundancy but no ambiguity
• Redundancy exists in the genetic code since
2 or more codons differing only in their
third base can code for the same amino acid
(UUU and UUC both code for
phenylalanine).
• Ambiguity is absent, since codons code for
only 1 amino acid.
The Correct Ordering
• The correct ordering and grouping of
nucleotides is important in the molecular
language of cells.
• Reading Frame=The correct grouping of
adjacent nucleotide triplets into codons that
are in the correct sequence on mRNA.
• The cell reads the message in the correct
frame as a series of non-overlapping 3-letter
words.
Common Genetic Language
• The genetic code is shared nearly
universally among living organisms.
• EX: The RNA codon CCG is translated into
proline in all organisms whose genetic
codes have been examined.
• Mitochondria and chloroplasts have their
own DNA codes for some proteins.
• Several ciliates depart from standard code;
codons UAA and UAG are not stop signals
but code for glutamine.
Transcription
• Transcription of mRNA from template DNA is
catalyzed by RNA polymerases, which:
– Separate the 2 DNA strands and link RNA nucleotides
as they base-pair along the DNA
– Add nucleotides only to the 3’ end
There are several types of RNA polymerase. Prokaryotes
have only 1 type of RNA polymerase but Eukaryotes
have 3 types, 1 for each type of RNA.
RNA polymerase II catalyzes synthesis mRNA.
Three Steps in Transcription
•
•
Transcription Unit—Nucleotide sequence on the
template strand of DNA that is transcribed into a
single RNA molecule; it includes the initiation
and termination sequences, as well as the
nucleotides in between.
Transcription occurs in 3 steps:
1.
2.
3.
Polymerase binding and initiation
Elongation
Termination
RNA polymerase Binding and
Initiation
• Promoter—region of DNA that
includes the site where RNA
polymerase binds and
transciption begins.
• In Eukaryotes, the promoter is
about 100 nucleotides long and
consists of the initiation site and
a few nucleotide sequences that
help initiate transcription.
• Transcription factors—DNAbinding proteins that bind to
specific DNA nucleotides at the
promoter that help RNA
polymerase recognize and bind
to the promoter region
TATA Box
• TATA box—a short
nucleotide sequence at the
promoter that is rich in
thymine and adenine,
about 25 nucleotides from
the initiation site.
• RNA polymerase II
recognizes the complex
between the bound TATA
transcription factor and
the DNA binding site
Elongation of the RNA Strand
• Once transcription begins,
RNA polymerase II moves
along DNA and:
1. Untwists and opens a short
segment of DNA exposing
about 10 nucleotides.
2. Links incoming RNA
nucleotides to the 3’ end of
the elongating strand.
During transcription, mRNA
grows about 30-60
nucleotides per second. As
it elongates, MRNA peels
away from the DNA
template.
Termination
• Transcription proceeds until RNA polymerase
reaches a termination site on DNA.
• Terminator sequence—DNA sequence that signals
RNA polymerase to stop transcription and to
release the RNA molecule and DNA template
• In Eukaryotes, the most common terminator
sequence is AATAAA.
• Prokaryotic mRNA is ready for translation
immediately; eukaryotic mRNA must be
processed before it leaves the nucleus and
becomes functional.
Translation is RNA-directed
synthesis of a polypeptide.
• Transfer RNA (tRNA) is the interpreter between
the mRNA and the amino acid sequence of the
polypeptide.
• tRNA aligns the appropriate amino acids to form a
new polypeptide. tRNA must:
– Transfer amino acids from the cytoplasm’s amino acid
pool to a ribosome.
– Recognize the correct codons in mRNA.
Molecules of tRNA are specific for only 1 particular
amino acid. Each type of tRNA associates a distinct
mRNA codon with one of the 20 amino acids.
ANTICODON
• One end of a tRNA
molecule attaches to a
specific amino acid.
• The other end attaches to
an mRNA codon by base
pairing with its anticodon.
• Anticodon—a nucleotide
triplet in tRNA that base
pairs with a
complementary triplet
(codon) in mRNA
Anticodon details:
• tRNAs decode the genetic message, codon
by codon.
• For ex: the mRNA codon UUU is
translated as the amino acid phenylalanine.
The tRNA that transfers phenylalanine to
the ribosome has an anticodon of AAA.
• As tRNAs deposit amino acids in the
correct order, ribosomal enzymes link them
into a chain.
Structure and Function of tRNA
•
•
•
•
•
•
•
All types of RNA are transcribed
from DNA.
tRNA must travel into the
cytoplasm where translation
occurs.
tRNA can be used repeatedly.
tRNA is a single-stranded RNA
about 80 nucleotides long.
There are 45 distinct types of
tRNA; some recognize 2 or 3 of the
mRNA codons.
Wobble—the ability of 1 tRNA to
recognize 2 or 3 different mRNA
codons; occurs when the 3rd base of
the tRNA anticodon has some play
or wobble, so that it can H-bond
with more than 1 kind of base in
the 3rd position of the codon.
Ex: The base U in the wobble
position of tRNA can pair with
either A or G in the codon.
A Unique Base
• Some tRNA molecules contain a modified
base called inosine (I), which is in the
wobble position and can base pair with U,
C, or A in the 3rd position of an mRNA.
• For example, a single tRNA with the
anticodon CCI will recognize 3 mRNA
codons: GGU, GGC, or GGA—all of
which code for glycine.
Aminoacyl-tRNA Synthetases
•
•
•
1.
2.
•
Aminoacyl-tRNA synthetase—a type of enzyme that
catalyzed the attachment of amino acid to its tRNA.
Each of the 20 amino acids has a specific aminoacyltRNA synthetase.
2 steps in attachment of an amino acid:
Activation of the amino acid with AMP. The
synthetase’s active site binds the amino acid and ATP;
the ATP loses 2 phosphate groups and attaches to the
amino acid as AMP.
Attachment of the amino acid to tRNA. The appropriate
tRNA covalently bonds to the amino acid, displacing
AMP from the enzyme’s active site.
The aminoacyl-tRNA complex releases from the enzyme
and transfers its amino acid to a growing polypeptide on
the ribosome.
Ribosomes
• Ribosomes have 2 subunits
(small and large) which are
separate when not involved
in protein synthesis.
• Ribosomes are composed
of about 60% ribosomal
RNA (rRNA) and 40%
protein.
• The large and small
subunits of eukaryotes are
constructed in the
nucleolus, move through
nuclear pores, and
assembled in the cytoplasm
only when attached to
mRNA.
A and P Sites
• Prokaryotic ribosomes are
smaller with different molecular
composition than eukaryotic.
• Antibiotics tetracycline and
streptomycin can be used to
combat bacterial infections
because they inhibit bacterial
protein synthesis but not
eukaryotic.
• In addition to a mRNA binding
site, each ribosome has 2 tRNA
binding sites (A and P).
• The P site (Peptidyle-tRNA)
holds the tRNA carrying the
growing polypeptide chain.
• The A site (Aminoacyl-tRNA)
holds the tRNA carrying the
next amino acid to be added.
Building A Polypeptide
•
1.
2.
3.
•
•
Translation occurs in 3 stages:
Initiation
Elongation
Termination
All 3 stages require enzymes and other protein
factors.
Initiation and elongation also require energy
provided by GTP—a molecule closely related to
ATP.
Initiation
• The assembly of the initiation
•
•
•
•
Initiation must bring together the
mRNA, the first amino acid attached to
its tRNA, and the 2 ribosomal subunits.
In eukaryotes, the small ribosomal
subunit binds first to an initiator tRNA
with the anticodon UAC and the amino
acid methionine.
The small ribosomal subunit next binds
to the 5’ end of mRNA.
The bound initiator tRNA finds and
base pairs with initiation or start codon
on mRNA, AUG.
•
•
•
•
complex—small ribosomal subunit,
initiator tRNA and mRNA—
requires:
– Protein initiation factors that
are bound to the small
ribosomal subunit.
– One GTP molecule to stabilize
the binding of initiation factors
and once hydrolyzed, drives
the attachment of the large
ribosomal subunit.
A large ribosomal subunit binds to
the small one to form a functional
ribosome.
Iniation factors attached to the
small subunit are released allowing
the large subunit to bind with the
small one.
The initiator tRNA fits into the P
site on the ribosome.
The vacant A site is ready for the
next aminoacyl-tRNA.
Elongation
•
Several proteins called elongation factors take part in
this 3-step process which adds amino acids one by one.
a.
b.
c.
Codon recognition. The mRNA codon in the A site forms
hydrogen bonds with the anticodon of an entering tRNA
carrying the next amino acid. An elongation factor directs tRNA
into the A site. Hydrolysis of GTP provides the energy.
Peptide bond formation. An enzyme, peptidyl transferase,
catalyzes the formation of a peptide bond between the
polypeptide in the P site and the new animo acid in the A site.
Peptidyl transferase is part of the large ribosomal subunit and
consists of ribosomal proteins and rRNA.
Translocation. The tRNA in the P site releases from the
ribosome, and the tRNA in the A site is translocated to the P site.
GTP hydrolysis provides energy for each translocation step.
Termination
• Termination codon (stop codon)—Base triplet
(codon) on mRNA that signals the end of
translation
• UAA, UAG, and UGA are stop codons and do not
code for an amino acid.
• When a stop codon reaches the ribosome’s A site,
a protein release factor binds to the codon and
initiates the hydrolyzes of the bond between the
completed polypeptide and the tRNA in the P site;
this frees the polypeptide and the tRNA; the 2
ribosomal subunits dissociate from mRNA and
separate into the small and large subunits.
Other Facts
• Polyribosomes—a cluster of ribosomes
simultaneously translating an mRNA
molecule.
• Prokaryotes lack nuclei, so transcription is
not segregated form translation and may
begin before transcription is complete.
• Eukaryotes process or modify mRNA
before it leaves the nucleus.
mRNA Modification
• 5’Cap—modified guanine nucleotide (GTP) that is
added to the 5’ end of mRNA shortly after
transcription begins; protects mRNA from
degradation and helps small ribosomal subunits
recognize the attachment site on mRNA’s 5’ end.
• Poly-A tail—sequence of about 200 adenine
nucleotides added to the 3’ end of mRNA before it
exits the nucleus
Gene Splicing
• The original transcript, or precursor mRNA, is
heterogeneous nuclear RNA (hnRNA); it is
processed before leaving the nucleus.
• Introns—noncoding sequences in DNA that
intervene between coding sequences (exons); are
initially transcribed but are not translated because
they are excised before mRNA leaves the nucleus
• Exons—coding sequences of a gene that are
transcribed and expressed.
• RNA splicing—RNA processing that removes
introns and joins exons from eukaryotic hnRNA to
produce mature mRNA
snRNPs
• Small nuclear ribonucleoproteins (snRNPs)complexes of proteins and small nuclear RNAs
that are found only in the nucleus; some
participate in RNA splicing.
– Composed of small nuclear RNA (snRNA) with less
than 300 nucleotides, and 7 or more proteins
Spliceosome—a large molecular complex that catalyzes
RNA splicing reactions; composed of snRNPs and
other proteins.
Ribozymes—RNA molecules that can catalyze reactions
by breaking and forming covalent bonds; rRNA
functions as an enzyme during translation
Mutation
• Mutation—a permanent change in DNA that
can involve large chromosomal regions or a
single nucleotide pair.
• Point mutation—a mutation limited to 1 or
2 nucleotides in a single gene
• Mutagenesis—the creation of mutations
• Mutagen—physical or chemical agents that
interact with DNA to produce mutations
– Radiation and certain chemicals including base
analogues can cause mutations.
Types of Point Mutation
• Base-pair substitution—the replacement of 1 base
pair with another; occurs when a nucleotide and its
partner from the complementary DNA strand are
replaced with another pair of nucleotides
according to base-pairing rules.
• Missense mutation—base-pair substitution that
alters an amino acid codon to a new codon that
codes for a different amino acid.
• Nonsense mutation—base-pair substitution that
changes an amino acid codon to a chain
termination codon, or vice versa
Types of Point Mutation
• Insertions or Deletions
• Base-pair insertion—the insertion of 1 or
more nucleotide pairs into a gene
• Base-pair deletion—the deletion of 1 or
more nucleotide pairs from a gene
• Frameshift mutation—a base-pair insertion
or deletion that causes a shift in the reading
frame, so that codons beyond the mutation
will be the wrong grouping of triplets and
will specify the wrong amino acids.