1. The genetic code is composed of nucleotide triplets called codons that specify individual amino acids. 2. Experiments confirmed that the genetic code is a triplet code and that each codon corresponds to a specific amino acid, with some codons coding for the same amino acid (degenerate). 3. Key properties of the genetic code include it being triplet-based, non-overlapping, unambiguous, degenerate, and nearly universal across organisms.

1. The Genetic Code
Objectives:
Understand the triplet nature of the genetic code, and know the meaning of the term codon.
Know that the code is degenerate, and what that means.
Know that the code is unambiguous, and what that means.
Know the identities of the start and stop codons, and understand how they work.
Genetic information is stored in DNA which was clear from the experiments of Avery, Macleod, and McCarty
and Hershey and Chase. However, these experiments did not explain how DNA stores genetic information.
Elucidation of the structure of DNA by Watson and Crick did not offer an obvious explanation of how the
information might be stored. DNA was constructed from nucleotides containing only four possible bases (A,
G, C, and T). The big question was: how do you code for all of the traits of an organism using only a four
letter alphabet?
Central dogma of molecular biology. The information stored in DNA is ultimately transferred to protein,
which is what gives cells and tissues their particular properties. Proteins are linear chains of amino acids,
and there are 20 amino acids found in proteins. So the real question becomes: how does a four letter
alphabet code for all possible combinations of 20 amino acids?
By constructing multi-letter "words" out of the four letters in the alphabet, it is possible to code for all of
the amino acids. Specifically, it is possible to make 64 different three letter words from just the four letters
of the genetic alphabet, which covers the 20 amino acids easily. This kind of reasoning led to the proposal
of a triplet genetic code.
Experiments involving in vitro translation of short synthetic RNAs eventually confirmed that the genetic
code is indeed a triplet code. The three-letter "words" of the genetic code are known as codons. This
experimental approach was also used to work out the relationship between individual codons and the
various amino acids. After this "cracking" of the genetic code, several properties of the genetic code became
apparent:

2. The Genetic Code

3. • The genetic code is composed of nucleotide triplets. In other words, three
nucleotides in mRNA (a codon) specify one amino acid in a protein.
• The code is non-overlapping. This means that successive triplets are read in order.
Each nucleotide is part of only one triplet codon.
• The genetic code is unambiguous. Each codon specifies a particular amino acid, and
only one amino acid. In other words, the codon ACG codes for the amino acid
threonine, and only threonine.
• The genetic code is degenerate. In contrast, each amino acid can be specified by
more than one codon.
• The code is nearly universal. Almost all organisms in nature (from bacteria to
humans) use exactly the same genetic code. The rare exceptions include some
changes in the code in mitochondria, and in a few protozoan species.
•Some of these properties will be examined in more detail.
A Non-overlapping Code The genetic code is read in groups (or "words") of three
nucleotides. After reading one triplet, the "reading frame" shifts over three letters,
not just one or two. In the following example, the code would not be read
GAC ACU CUG UGA
Rather, the code would be read
GAC, UGA, CUG, ACU...
GAC UGA CUG ACU...

4. Degeneracy of the Genetic Code
There are 64 different triplet codons, and only 20 amino acids. Unless some
amino acids are specified by more than one codon, some codons would be
completely meaningless. Therefore, some redundancy is built into the
system: some amino acids are coded for by multiple codons. In some cases,
the redundant codons are related to each other by sequence; for example,
leucine is specified by the codons CUU, CUA, CUC, and CUG. Note how the
codons are the same except for the third nucleotide position. This third
position is known as the "wobble" position of the codon. This is because in
a number of cases, the identity of the base at the third position can wobble,
and the same amino acid will still be specified. This property allows some
protection against mutation - if a mutation occurs at the third position of a
codon, there is a good chance that the amino acid specified in the encoded
protein won't change.

5. Reading Frames
Because the genetic code is triplet based, there are three possible ways
a particular message can be read, as shown in the following figure:
Clearly, each of these would yield completely different results.
To illustrate the point using an analogy, consider the following set of
letters: theredfoxatethehotdog
If this string of letters is read three letters at a time, there is one reading
frame that works: the red fox ate the hot dog
and two reading frames that produce nonsense:
t her edf oxa tet heh otd og
th ere dfo xat eth eho tdo g
Genetic messages work much the same way: there is one reading frame
that makes sense, and two reading frames that are nonsense

6. So how is the reading frame chosen for a particular mRNA?
The answer is found in the genetic code itself. The code contains
signals for starting and stopping translation of the code. The start
codon is AUG. AUG also codes for the amino acid methionine, but the
first AUG encountered signals for translation to begin. The start codon
sets the reading frame: AUG is the first triplet, and subsequent triplets
are read in the same reading frame. Translation continues until a stop
codon is encountered. There are three stop codons: UAA, UAG, and
UGA. To be recognized as a stop codon, the triplet must be in the same
reading frame as the start codon. A reading frame between a start codon
and an in-frame stop codon is called an open reading frame. Let's see
how a sequence would be translated by considering the following
sequence:
5'-GUCCCGUGAUGCCGAGUUGGAGUCGAUAACUCAGAAU-3‘
First, the code is read in a 5' to 3' direction. The first AUG read in that
direction sets the reading frame, and subsequent codons are read in
frame, until the stop codon, UAA, is encountered.

7. 5'-GUCCCGUGAUGCCGAGUUGGAGUCGAUAACUCAGAAU-3
Met Pro Ser Trp Ser Arg Stop
In this sequence, there are nucleotides at either end that are outside of
the open reading frame. Because they are outside of the open reading
frame, these nucleotides are not used to code for amino acids. This is a
common situation in mRNA molecules. The region at the 5' end that is
not translated is called the 5' untranslated region, or 5' UTR. The region
at the 3' end is called the 3' UTR. These sequences, even though they do
not encode any polypeptide sequence, are not wasted: in eukaryotes
these regions typically contain regulatory sequences that can affect
when a message gets translated, where in a cell an mRNA is localized,
and how long an mRNA lasts in a cell before it is destroyed

9. Through the experiments it has been proved that the mRNA codons of the
genetic code have the following properties :
The code is triplet : Triplet code consists of 4x4x4 = 66 codons may code
for 20 essential amino acids. The triple code of mRNA has been accepted.
The code is degenerate : There are 64 codons in the genetic code for 20
amino acids of which 4 codons are the signals. Therefore, 61 codons are to
code for amino acids.It means that more than one codon may be coding
for individual amino acid.
The code is non-overlapping : The genetic code is non-overlapping which
means that the same letter does not take part in the formation of more than
one codon.
The code is non-ambiguous : A particular codon will always code for the
same amino acid. It may also be that the same amino acid may be coded
by two different codons. However, when one codon codes for two amino
acids, it is called ambiguous.

10. 1.All 64 codons are used. 61 of them can be assigned to certain
amino acids, the other three are stop signals. One of the codons can
act both as an amino acid codon and as a start signal.
2.The different amino acids have different numbers of accompanying
codons. For some, like Met or Trp exists just one codon, for others
two or four and for some (Ser, Arg) even six. The frequency of the
codons and the frequency of their amino acid is correlated. An
exception is Arg, that has six codons but is underrated regarding its
frequency in proteins.
3.The codons are not assigned randomly. The first two nucleotides of
a codon have a higher informational value than the third one, GUU,
GUC, GUA and GUG, for example, do all encode Val. Codons rich in
UC encode hydrophobic, such rich in AG hydrophilic amino acids.

11. Many (nearly 30%) of all base substitutions do not change the encoding
properties, for example:
UUU > UUC: Phe > Phe
Even if a base substitution causes an amino acid exchange is the
chemical character of the side chain in most cases conserved
(conservative exchanges). The genetic code can consequently be
regarded as extremely conservative:
UUU > UUG: Phe > Leu
CUC > AUC: Leu > Ile
AAA > AGA: Lys+ > Arg+
AAA > GAA: Lys+ > Asp-
Exceptions exist (radical exchanges):
GAG > GUG: Glu- > Val
GAA> GUA: Glu- > Val

12. The code is comma less : The genetic code is without comma i.e. no
punctuations are required between the two codons. There are no
demarcating signals between two codons. This result is continuous
coding of amino acid without interruption . No codons are left uncoded
which will be like UUUCUCGUAUCC.
The code has polarity : The code has polarity which is read between the
fixed start and stop codons. The start codon is also known as initiation
codon, and stop codon as termination codon.
The code is Universal : Though the genetic code has been worked out by
using in vitro systems of microorganisms, yet there is no doubt of being
its universal for all groups of micro organisms.

13. Different organisms exhibit different statistical preferences of triplet codon
usage, as well as using the amino acids in widely varying proportions.
See Of URFs and ORFs' by Russell Doolittle, University Science Books
(1986) ISBN 0-935702-54-7.
The Mitochondrial Genetic Code
Human mitochondrial DNA encodes only 22 tRNA species and these are
the only tRNAs used for the translation of mitochondrial mRNAs. This is
accomplished by an extreme form of wobble in which U of the anticodon in
tRNA can pair with any of the four bases in the third codon position of the
mRNA, allowing four codons to be recognized by a single tRNA. In addition
some codons specify different amino acids in mitochondria than in the
universal code.
Differences between the Universal and Mitochondrial Genetic Codes
Human
Codon Universal code
mitochondrial code
UGA Stop Trp
AGA Arg Stop
AGG Arg Stop
AUA Ile Met

14. The "Wobble" Hypothesis
Even before the genetic code had been elucidated, Francis Crick
postulated that base pairing of the mRNA codons with the tRNA
anticodons would require precision in the first two nucleotide positions
but not so in the third position (the precise conformation of base pairs,
which refers to the hydrogen bonding between A-T (A-U in RNA) and C-G
pairs is known as Watson-Crick base pairing). The third position, in
general, would need to be only a purine (A or G) or a pyrimidine (C or U).
Crick called this phenomenon "wobble."
This less-than-precise base pairing would require fewer tRNA species.
For example, tRNAGlu could pair with either GAA or GAG codons. In
looking at the codon table, one can see that, for the most part, the first
two letters are important to specify the particular amino acid. The only
exceptions are AUG (Met) and UGG (Trp) which, as indicated above, have
only one codon each.

15. The Wobble Hypothesis - 1966, Francis Crick
The genetic code is degenerate: one amino acid may be encoded by several
different codons.
unmixed codon families - first two bases always code for the same amino
acid (there's practically no need to read the third base of the codon) e.g. - leu,
val, ser, rpo, thr, ala etc.
mixed codon families - first two bases may be included in the code for more
than one different amino acid or for an amino acid and a start or stop codon.

16. Crick termed this redundancy "wobble": the code was not rigid, and there
was room for error.
Similar codons code for amino acids with similar physical properties. For
example:
a "U" in the center position always encodes a hydrophobic aa. A mutation at
the two outer positions will not change that.
negatively charged aa's (e.g. aspartate, glutamate) always begin with GA. A
mutation in the 3rd position will not change that.
This is evidence that a triplet code not only allows for more diversity, but
also provides a margin of error in terms of deleterious mutations.

18. EXCEPTIONS TO THE UNIVERSAL GENETIC CODE
Organism Normal codon Usual meaning New meaning
Mammalian AGA, AGG Arginine Stop codon
mitochondria AUA Isoleucine Methionine
UGA Stop codon Tryptophan
Drosophila AGA, AGG Arginine Serine
mitochondria AUA Isoleucine Methionine
UGA Stop codon Tryptophan
Yeast AUA Isoleucine Methionine
mitochondria UGA Stop codon Tryptophan
CUA, CUC, CUG,
Leucine Threonine
CUU
Higher plant UGA Stop codon Tryptophan
mitochondria CGG Arginine Tryptophan
Protozoan nuclei UAA, UAG Stop codons Glutamine
Mycoplasma
capricolum UGA Stop codon Tryptophan
bacteria