The Genetic Code

1
The Genetic Code

• Dr. Jason Linville
• University of Alabama at Birmingham
• jglinvil_at_uab.edu

2
Summary

• Structure of proteins
• Cracking the genetic code

How does DNA code for proteins?
3
Structure of Proteins

• Proteins are polymers of amino acids
• Joined by peptide bonds

AGC TAG CTT ATA CTC TAT CTC TTT
Amino Acid
Amino Acid
Amino Acid
Amino Acid
Amino Acid
Amino Acid
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Protein Structure gt Amino Acids

• Amino Acids

• 20 amino acids found in proteins
• All amino acids have the same general structure

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Protein Structure gt Amino Acids

• Amino Acids

Each central carbon atom (a-C) has

• Hydrogen atom
• Carboxyl group

• Amino group
• R group

The R group is different for each amino acid.
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Protein Structure gt Amino Acids

• Amino Acids

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Protein Structure gt Amino Acids

• Non-Polar
• alanine, valine, leucine, isoleucine, proline,
  methionine, phenylalanine, tryptophan

• Uncharged Polar
• glycine, serine, threonine, cysteine, asparagine,
  glutamine, tyrosine

• Positive Polar
• Lysine, arginine, histidine

• Negative Polar
• Aspartic acid, glutamic acid

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Protein Structure gt Peptide Bond

• Peptide Bond

• Amino group reacts with carboxyl group
• Peptide bond H2O
• Protein has an amino end and carboxyl end

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Protein Structure gt Peptide Bond

• Peptide Bond

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Protein Structure gt Levels

• Levels of Protein Structure

• Primary Structure
• Secondary Structure
• Tertiary Structure
• Quaternary Structure

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Protein Structure gt Levels

• Primary Structure

• The sequence of amino acids

• This sequence alone will specify the secondary,
  tertiary, and quaternary structures (sort of).
• Environmental conditions also affect structure

12
Protein Structure gt Levels

• Secondary Structure

• The repeating configuration taken up by the amino
  acid chain.
• a-helix
• ß-sheet

• Both are stabilized by hydrogen bonds
• Proteins are composed of many a-helices and
  ß-sheets, and unorganized regions

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Protein Structure gt Levels

• Tertiary Structure

3-D formation of the polypeptide

• Held together with
• Hydrogen bonds
• Disulphydryl bridges
• Hydrophobic/hydrophilic interactions

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Protein Structure gt Levels

• Tertiary Structure

Disulphydryl Bridges form between two cysteine
residues.
Hydrophobic/Hydrophilic interactions refers to
tendency of hydrophobic residues (non-polar) to
be shielded from water (interior of protein).
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Protein Structure gt Levels

• Quaternary Structure

Orientation of 2 or more polypeptides to form one
protein.
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Protein Structure gt Amino Acids

• Overall Structure

• Primary structure specifies the secondary,
  tertiary, and quaternary structures.

• This is understood to such a degree that given
  the gene sequence, secondary structures can be
  predicted using computer software.

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Protein Structure gt Amino Acids

• Overall Structure

• Tertiary and Quaternary structures are more
  difficult to predict.

• If a protein is denatured (heat or pH), it can
  spontaneously renature to form the same tertiary
  and quaternary structures.

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Cracking the Code

• Early Work

• Genes and Proteins are colinear
• Order of nucleotides in gene directly relates to
  the order of amino acids in protein
• 5 end of gene is amino-terminus of protein

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Cracking the Code

• Early Work

• Triplet of nucleotides 1 amino acid
• Single or doublet wouldnt be able to code for
  the 20 amino acids.
• Triplet provides 64 code words for 20 amino acids

How was this determined?
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Cracking the Code

• Identifying triplets (experiment)

• Inserted single base pairs into unimportant
  section of DNA gene
• One insertion nonsense
• Two insertions nonsense
• Three insertions sense returns (with one extra
  amino acid in unimportant region)

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Cracking the Code

• Important experiments

• Protein synthesis with homopolymers
• Synthesis with random heteropolymers
• Synthesis with ordered heteropolymers
• Triplet binding assay

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Cracking the Code

• Earlier work

• Synthesis of artificial RNA molecules
• Polynucleotide phosphorylase synthesizes RNA in
  the absence of DNA template
• Can make RNA with known sequence

• Cell-free protein synthesis
• cell-free (in vitro) system can synthesize
  proteins from RNA molecule

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Cracking the Code

• Homopolymers

• Example 5 UUUUUUUUUU 3

• Made from one type of nucleotide

• Poly U resulted in protein comprised of only
  phenylalanine

• UUU phenylalanine

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Cracking the Code

• Random Heteropolymers

• Example 5 ACCACACCAAAC 3

• Made from two nucleotides

• Eight different codons possible
• (AAA,AAC,ACA,ACC,CAA,CCA,CAC,CCC)

• Six different amino acids result
• Which codon codes for which amino acid?

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Cracking the Code

• Random Heteropolymers

• Problem solved by adding nucleotides in different
  ratios, then measuring relative amounts of amino
  acids

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Cracking the Code

• Ordered Heteropolymers

• Polymerize dinucleotides or trinucleotides

• Dinucleotide (AC) polymer can be read two ways
• ACA or CAC

• Trinucleotide (UGU) polymer can be read three
  ways
• UGU, GUU, UUG

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Cracking the Code

• Triplet Binding Assay

• Mix known triplet (ACU) with tRNA
• One type of AA is radiolabeled, all others are
  not.
• Only the tRNA (with specific AA) that matches the
  triplet will bind to ribosome
• Isolating and testing ribosomes for radiolabel
  will tell if triplet and AA match

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Cracking the Code
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Cracking the Code

• Features of code

• Degenerate more than one codon can code for a
  single amino acid
• Punctuation codons some codons code for
  terminating translation
• Not universal May change in mitochondrial genes
  or among species