The Genetic Code - AGHAMAZING...
Transcript of The Genetic Code - AGHAMAZING...
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Translating the Genetic Code
DANILO V. ROGAYAN JR.Faculty, Department of Natural Sciences
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Figure 13.2
An overview of gene expression
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The Idea of A Code
• 20 amino acids
• 4 nucleotides
• How do nucleic acids
composed of 4
nucleotides specify the
synthesis of proteins
composed of 20 different
amino acids?
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The Coding Problem
• 1-1 correspondence – NO – 4 possibilities
• 2-1 correspondence – NO – 16 possibilities
• 3-1 correspondence – YES – 64
possibilities
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Determination of Triplet Nature of Code
• Aacridines & flavinoids cause single nucleotide deletions and insertions respectively
• Insertion of 1 base shifts frame & creates non-sense mutation
• Deletion of base in mutant restores frame (suppresses 1st
mutation)
• Recombination between mutants showed that insertion of 1 or 2 bases or removal of 1 or 2 bases doesn't restore frame, but insertion of 3 or removal of 3 allows frame to continue
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Crick’s Experiments to Determine
Triplet Nature of Code
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So.. The code is 3 letter words, but what
about punctuation?
GROWANDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEW
• There is a message, but one must start at the right place to read it
• Code written in three letter words - codon
• There are three reading frames, but only one gives an intelligible message – frame b– NOW THE CAT SAW THE DOG BUT DID NOT RUN
• A start codon (NOW) and a stop codon (END) define the frame to use
a
b
c
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Any frame is potentially the
reading frame!
OWANDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEWa
b
c
ROWANDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEWa
b
c
The actual reading frame is called the Open Reading Frame or ORF
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Reading Frames & Mutation
Types• Frame shift mutations
– Original reading frame is frame a
– Insertions or deletions shift the reading frame
ROWANDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEWa
b
c
ROWNDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEWa
b
c
^
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Reading Frames & Mutations
• Once a ribosome begins translation in a particular
frame (a) it does not shift frames
• Therefore, if a mutation shifts the reading frame in
the mRNA, the ribosome will read the wrong
frame.
ROWANDNOWTHECATSAWTHEDOGBUTDIDNOTRUNENDSEWa
b
c
ROWANDNOWTHECATSAWTHEADOGBUTDIDNOTRUNENDSEWa
b
c
^
NOW THE CAT SAW THE ADO GBU TDI DNO TRU NEN DSE W..
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Deciphering the Code
• Each amino acid in a protein is specified by 3
nucleotides of codon
• Each codon specifies only ONE amimo acid
• There are 64 possible codons but only 20
amino acids
• Degeneracy
– An amino acid can be specified by multiple codons
– A given codon still only specifies only one amino
acid
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Deciphering the Code: Three
Approaches• Synthesis and translation of
homopolymer RNA & ratio polymer RNA
followed by amino acid analysis
• Synthesis and translation of di, tri and
tetra nucleotide repetitive RNA polymers
followed by amino acid analysis
• Triplet RNA-tRNA binding assay and
amino acid analysis
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Synthetic RNA Templates and In Vitro
Translation
• Synthesis of homopolymeric RNAs– UUUUUUUUUU, AAAAAAAAAA, CCCCCCCCC, &
GGGGGGGGG
• When translated produced polypeptides– poly phenylalanine (UUU), polyproline (CCC), polylysine
(AAA), and polyglycine (GGG)
• Therefore 4 codons were determined– UUU = phe
– CCC = pro
– AAA = lys
– GGG = gly
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• The enzyme polynucleotide phosphorylase– polymerizes ribonucleoside diphosphates (NDPs) into RNA
– It does not use a template, the order of nucleotides is random
– The nucleotide sequence is controlled by the relative abundance of NDPs
• For example, if 70% GDP and 30% UDP are mixed together, then …
Possible
Codons
Percentage in the Random
Polymer
GGG 0.7 x 0.7 x 0.7 = 0.34 = 34%
GGU 0.7 x 0.7 x 0.3 = 0.15 = 15%
GUU 0.7 x 0.3 x 0.3 = 0.06 = 6%
UUU 0.3 x 0.3 x 0.3 = 0.03 = 3%
UGG 0.3 x 0.7 x 0.7 = 0.15 = 15%
UUG 0.3 x 0.3 x 0.7 = 0.06 = 6%
UGU 0.3 x 0.7 x 0.3 = 0.06 = 6%
GUG 0.7 x 0.3 x 0.7 = 0.15 = 15%
Decoding the Genetic Code
Radiolabeled Amino Acid Added
(%) Amino Acid
Incorporated
Glycine (GGG + GGU, UGG, GUG)
49
(34 + 15)
Valine
(GGU, UGG, GUG, GUU, UUG, UGU)
21
(15 + 6)
Tryptophan(UGG, GUG, GGU)
15
Cysteine(UUG, GUU, UGU)
6
Leucine
(UUG, GUU, UGU)
6
Phenylalanine
(UUU)
3
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Polymers of Nucleotide Repeats
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Trinucleotide-tRNA Binding Analysis
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• Special codons:
– AUG (which specifies methionine) = start codon
• AUG specifies internal methionines also
– UAA, UAG and UGA = termination, or stop, codons
• The code is degenerate
– More than one codon can specify the same amino acid
• For example: GGU, GGC, GGA and GGG all code for lysine
– In most instances, the third base is the degenerate base
• It is sometime referred to as the wobble base
• The code is nearly universal
– Only a few rare exceptions have been noted
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Lakô hã salamát!Maraming salamat!