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DNA Transcription and Translation
Once we had discovered enzymes, we thought they were the most important thing so
the idea of one gene = one enzyme. Then realized it was proteins not enzymes, so
one gene = one protein. Upon further research, found that the protein that makes
up haemoglobin is controlled by two genes, not one. Finally, one gene = one
polypeptide.
One gene = one polypeptide. But now it is not good enough, many exceptions. But how
are polypeptides different? In their primary structure, they differ based on the amino
acids - which ones are used and how they are bonded together. There are 20 different
amino acids and they are used to make polypeptides. ( HL -remember, some are polar
and some are non-polar) So, the DNA molecule codes for the amino acids, which ones
and where. But the only variable thing about the DNA molecule is the nitrogen bases.
How can 4 bases or combination there of, code for 20 different amino acids?
1. if one base coded for 1 amino acid = 41 different amino acids coded.
2. if two bases coded for 1 amino acid = 42 different amino acids or 16 coded
3. if three bases coded for 1 amino acid = 43 different amino acids or 64 coded
Large gap between two and 3. Concluded that 3 nitrogen bases code for 1 amino acid.
But 64 is larger than 20 so some amino acids are coded for by more than one 3 base code
(codons)
So, it takes 3 nitrogen bases or 3 nucleotides to code for a specific amino acid – just like
it may take 3 letters to make up a word. A triplet code. But it also matters how the bases
are read, what order, where to start.
Our ability to extract the intended message or code depends on reading the bases in the
correct sequence or grouping.
The red dog ate the cat
If out of sequence =
The red cat ate the dog
Analogy – airline uses a three
Wrong grouping =
code for all the airports in the
There ddo gate the cat
world, area code for telephones
Start at the wrong spot =
uses 3 numbers
Her edd oga tet hec at
But how does the information that is encoded in the DNA actually get to the site of
protein synthesis – ribosomes? We can’t just let the DNA wander over there and unzip
itself. (prokaryotes can be we can’t) And its also too big to be leaving every time a
protein needs to be made. We need a messenger to read the base message and take it to
the ribosomes to be made. Enter the RNA.
“The DNA code is inert, in the same way that valuable encyclopaedias are stored safely
in libraries, rather than being consulted in factories. For daily use the cell relies on
disposable photocopies – RNA”. Power, Sex, Suicide. Mitochondria and the Meaning of Life by
Nick Lane
Three major differences between RNA and DNA. RNA is only one strand, the 5-carbon
sugar is ribose and it does not have the thymine base. Instead, it uses uracil. (bonds the
same way as thymine)
Know the structure of ribose and be able to compare to glucose, ribose has 5
carbons and glucose has 6 - in multiple choice
For HL
Transcribe – to copy from one form to another
Translate – to convey from one form to another, to convey the meaning; translate DNA
code into amino acid sequence.
To get from DNA to correct amino acid sequence involves two main steps; 1)
transcription – synthesis of RNA under the direction of DNA. They use the same
language (base pairs) so it is simply copied or transcribed.
2) translation – synthesis of a polypeptide. Translating the base code (called codon) into
the correct amino acid sequence.
Transcription
The first RNA used is messenger RNA – mRNA. It goes into the nucleus, through the
nuclear pores in the nuclear membrane,(remember the draw the pores in a drawing) and
reads the DNA. It is complementary rather than identical. And U instead of T.
It takes 3 base pairs or nucleotides to code for a specific amino acid – a triplet code.
Three bases on mRNA coding for an amino acid is called a codon. Looking at the
mRNA amino acid chart, we can see that some codons code for the same amino acid. Eg.
GAA and GAG both code for glutamic. This is called degeneracy – more than one code
for a single amino acid. Also, some codons do not code for an amino acid but regulate
processes like start and stop codons. There maybe degeneracy in the codons but there is
no ambiguity. That is, you may have two codons that code for the same amino acid but
you DO NOT have one codon that codes for two different amino acids. (I called
degeneracy – redundancy sometimes)
Universality – the genetic code is universal, that is all organisms use the same base codes
to code for the same amino acids. (but what they do with the amino acids is different.
Remember the 4 levels of proteins, amino acid sequence is the first level only). If you
know the right code sequence, you can make the same protein in any organism.
Important application – the making of insulin and factor VIII (for clotting). We know
the sequence for these proteins and have inserted it into the DNA of yeast cells. The
yeast will produce it in large quantities at a fast and easy rate.
Use zipper to show what RNA polymerase does
RNA polymerase is a large molecule that latches on and unwinds DNA and catalyzes the
reaction that attaches complementary nucleotides together to make the mRNA. The
polymerase moves down the DNA, untwisting it and exposing DNA. It can do 60
nucleotides/sec. When it reaches the termination region, there is a stop codon in it,
which will cause the RNA polymerase to unlatch and the newly made mRNA peels away.
The usual sequence is that DNA makes the RNA - except for retro viruses of which HIV
is one.. These type of viruses are mostly RNA and have an enzyme called
retrotranscriptase, that can make DNA from their RNA – going backwards or retro. This
new DNA is inserted into our (the host) genome and replicated every time we replicate
our DNA
HL - Yes more
So mRNA needs to ‘read’ the DNA. And not both sides, only one side or strand. The
DNA strand being transcribed is called the antisense strand or template. The mRNA will
be complementary to the antisense strand. The other strand is called the sense strand.
(don’t look at me, I didn’t make this words up). MAKE SURE you understand this.
RNA polymerase will act just like DNA polymerase. But how does it know where to
start? Only a part of the antisense strand needs to be read or transcribed. As you only
need one gene, not all the genes found on the DNA. Enter the promoter.
The promoter is a region of the DNA where RNA polymerase will attach to start
transcription. The promoter contains a start codon and other nucleotides. (It’s TAC on
the DNA or AUG complementary for RNA).
RNA polymerase can only work in a 53 direction which means it will attach to the
35 strand of the DNA so that it can work in a 5 3 direction. Make sure you can look
at a transcribing process and know the direction it is going.
When the RNA nucleotides show up to be attached, they show up as ribonucleoside
triphosphates. They will attach to each other by a hydrogen bond (covalent) and the 2
phosphate groups will be removed to supply the energy to do this.
The mRNA needs to be modified before it leaves the nucleus to head for the ribosomes.
Eukaryotic cells have our genes interspersed with large, non-coding pieces of repetitive
sequences. These bases need to be cut out, leaving only the coding parts. The noncoding parts, on the mRNA, are called introns and the coding parts are called exons
(because they are expressed). The process is called splicing and of course, enzymes do it.
Once introns are removed, it is called a mature RNA. Also adds a cap to 5’ end and a tail
to 3’. Cap acts like an “attach here” sign and tail protects.
Prokaryotes do not have genes with pieces of junk coding in them so they do not have to
go through this process. (We don’t sound so special anymore). Introns are translated but
NOT made into polypeptides
Introns not useless. Maybe controlling expression of genes, could allow
recombination or increase crossing over probability
Big question is how does DNA know where to unpack, which specific site is needed
for transcription? Environmental, hormonal, negative feedback lac operon type
or??? Epigenetics
Translation – codons translated into amino acids.
DNA and proteins don’t speak the same language so RNA acts as interpreter. RNA is
assembled and disassembled as needed from spare parts found in our cells Amino acts
like a 2 prong plug-in, mRNA like a 3 prong outlet and tRNA is the adapter to let
both fit together
The cell needs to interpret the genetic message and build a protein according to the
message. The message is a series of codons on mRNA and must be translated by tRNA.
The tRNA is specific, carrying one specific amino acid with them. By reading the codon,
- a complementary base pairing, the right tRNA with the right amino acid locks in and the
polypeptide is started. The cytoplasm has a good supply of the amino acids, either by
synthesizing them or taking from the blood. (Essential amino acids are ones we can’t
make and must eat)
The ribosome binds to the mRNA and moves along it until it reaches the start codon. As
the tRNA arrives at the ribosome, it has its specific amino acid attached at one end. At
the other end is a base triplet called an anticodon – complementary to the codon. The
tRNA plugs in by a hydrogen bond. tRNA deposits amino acids in the order prescribed
by mRNA codon and ribosomal enzymes join the amino acids together to form a peptide.
With a peptide bond.
Again HL
mRNA is complementary to the antisense DNA. tRNA is complementary to mRNA. So
tRNA is the same as antisense DNA and mRNA is the same as sense DNA – except for T
and U.
The structure of ribosomes. Ribosomes supply the place where translation happens. It is
the garage, which has all the tools to make the car. It is made up of a large and small
unit. Both units contain rRNA and some proteins. (60% of ribosomes are rRNA). The
two units lock in and supply and slot for the rRNA and bays for the tRNA.
Ribosomes are measured in a unit called “s”. (Suedberg number) Eukaryotes have
ribosomes that are 80 s while prokaryotes are 70 s. (There are ribosomes in the
mitochondria and chloroplasts – remember the endosymbiotic theory)
Prokaryotes ribosomes are chemically different. Medically, this is great as certain
antibiotics can paralysis prokaryotes ribosomes without harming Eukaryotes.
Tetracycline and streptomycin work this way. Transcription and translation are coupled
in prokaryotes as they happen together – due to no specific nuclear region or introns.
The structure of tRNA. Composed of one chain of about 80 nucleotides that have folded
into a cloverleaf shape with a 3 and a 5 end. The 3 end is the amino acid attachment and
it has the code of CCA. (or ACC) See Campbell page 305. Great pictures. Each amino
acid has a specific tRNA-activating enzyme that attaches the amino acid to its tRNA.
Note, some amino acids have more than one tRNA that can pick it up – degeneracy. The
energy required to attach it is supplied by ATP and the amino acid joins by a
condensation reaction – water is formed.
Look at page 306 of Campbell – figure 17.13 of enzymes joining the two together
Translation is divided into three parts – initiation, elongation and termination.
In initiation, mRNA binds to the small sub-unit of the ribosome. The ribosome then
slides along the mRNA until it reaches the start codon – AUG. tRNA and the large subunit are brought together. The anticodon of the tRNA joins with the complementary base
pair of the codon of the mRNA. The tRNA anticodon first joins with the start codon on
the mRNA and forms a hydrogen bond between the two.
The large sub-unit of the ribosome has two binding sites (like 2 bays) and a third exit site.
One called A for amino acid site, one called P for peptide site and the third is E for exit
site. P is for the tRNA that is in the process of reading and binding to mRNA and A is
for the next one in line. There is one binding site for mRNA. SEE DIAGRAMS.
Proteins called initiation factor help to do this. Energy is needed in the form of GTP
(guanosine triphosphate) which is changed to GDP. You also have an initiator tRNA
anticodon, which hooks in first and gets the whole ball rolling.
In elongation, the hydrogen bond between the anticodon and codon moves the mRNA
along. Kind of like gears locking into each other and turning or moving each other. A
second tRNA pairs with the next codon in the next binding site. The amino acid from the
tRNA in site A is transferred to the amino acid in the P site and a peptide bond is formed.
The tRNA that ‘lost’ its amino acid moves to the exit site and everyone shuffles over.
OF COURSE, it moves in a 53 direction on the mRNA. The tRNA in the exit site
leaves (to get another amino acid specific to its base sequence) and another tRNA pairs
with the next codon in the A site.
In termination, the stop codon is reached on the mRNA. (the stop or termination codon
stops the process by adding water to the end and not an amino acid) Once the
ribosome reaches the stop codon, the polypeptide is released – mRNA detaches. And the
ribosome splits into its small and large units. Once the polypeptide is made it starts to
fold to give its secondary or tertiary shape. Genes gave it the primary shape – that is the
sequence of amino acids and will also determine what shape it will form.
Free ribosomes synthesize proteins for use in the cell while rough or membrane bound
ribosomes synthesis proteins for export outside the cell ( Golgi, vesicles etc).
Ribosomes can make an average sized polypeptide in less than a minute. Several
ribosomes can be working on the same mRNA as many proteins are repeating sequences.
This can be seen under a light microscope. As one tRNA gets past the initiation site and
the start codon, another ribosome can attach. This cluster is called polyribosomes or
polysomes.
Mutations
Textbook – read p.68-69. Note mc. On page 70.
Mutations – inheritable mistake(s) in the genetic message. Important if it happens to the sex
cells. Low rates of mutation are good for variations in the species – natural selection. High rates
are harmful. Occasional mutations can lead to improved characteristics of novel capabilities but
more often than not, they are detrimental.
Mutagenic agents
Radiation – high-energy radiation like x-rays and gamma rays are highly mutagenic. When
radiation reaches the cell, the atoms absorb it and the electrons in the outer shell leave. Called
free radicals. Free radicals are highly chemically reactive and react with other cells and DNA.
Ultraviolet light – in DNA, the T and C bases absorb the energy and create free radicals
Chemicals All can cause a gene or point mutation, which can lead to a frame shift.
Gene mutation - a change in the base sequence of a gene. There are many types but main types
are a) base substitution b) deletion c) insertion d) inversion.
*Base substitution – substituting one base for another.
- new allele produced; as new amino acid will be coded for
- mRNA transcription will be affected; polypeptide will be different
- phenotype may be different; genotype will be different
- *Sickle-cell anaemia is an example –has 250000 bases and one is misspelled.
- *From GAG to GTG; structure of RBC different * must know name of bases - At low[CO2] they link together due to their shape glutamic acid replaced by valine
- So RBC shaped distorted and ability to pick up oxygen.
- Gives malaria resistance; symptoms
Deletion – where one base is deleted, eg. Cystic fibrous.
Inversion – 2 nucleotides invert
Analogy consider the faulty information conveyed in a sentence with which one single letter is
mutated
Intended = Susan is now arriving by air from New York
Substitution = Susan is not arriving by air from New York
Intended = Please say where you are
Insertion = Please stay where you are
Intended = I will send a friend to collect you
Deletion = I will send a fein dt oco lle cty ou
Insertion and deletion can sometimes be worse as it can lead to a frameshift. In a frameshift the
results are more dramatic as more bases are disrupted. Don’t forget, you can also get
chromosome mutations like nondisjunction
In base pair mutation → doesn’t always cause any change as there is degeneracy so still get the
same amino acid or if a different amino acid is coded, the protein is so big it might not make a
difference. BUT if it is the amino acids that are used for the active site of an enzyme → big
trouble.
You have many tumour-suppressing genes but as you get older, you pick up more mutations and
they accumulate. Or inherit faulty suppressor genes or something in the environment affects its.
One important one is p53 gene. Damage to DNA causes it to be stimulate which →halts cell
cycle allowing time to repair DNA, → turns on another group of genes that repair DNA and can,
if necessary cause → apoptosis or cell death so mutation not passed on.
Oncogenes – (onco is Greek for tumour) cancer-causing genes.