GENE to PROTEIN

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GENE to PROTEIN
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• Garrod (1909) hypothesized that the symptoms of
  an inherited disease reflect a persons inability
  to make a particular enzyme.
• The breakthrough in demonstrating the
  relationship between genes and enzymes when
  Beadle and Tatum began to search for mutants of
  bread mold. They discovered that mutants differ
  from wild type in their nutritional needs.
• Nutritional mutants are called auxotrophs.
• Beadle and Tatum were able to demonstrate the
  relationship between genes and enzymes by
  studying mutant forms of bread mold.

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Each gene dictates the production of one enzyme.
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• Many genes are made from two or more polypeptide
  chains therefore it is really one gene one
  polypeptide.
• Genes are typically hundreds or thousands of
  nucleotides long. The nucleic acids and proteins
  are written in two different languages. DNA must
  be converted to protein language.
• This is accomplished through the processes of
• Transcription
• Translation

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Prokaryotes vs Eukaroytes

• Prokaryotes lack nuclei and DNA is not segregated
  from ribosomes and transcription and translation
  occur in rapid succession.
• Eukaryotes have a nuclear envelop envelop and
  that segregates transcription in the nucleus from
  translation that occurs in the cytoplasm.

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• The flow of information from a gene to a protein
  is based a a triplet code.
• These three nucleotide words are codons.

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Cracking the Genetic Code

• Nirenberg 1961 NIH
• First codon decipher was UUU
• There are 64 codons
• A codon codes for only 1 amino acid
• The genetic code must have evolved very early in
  the history of life because it is nearly
  universal among living organisms.

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Transcription

• Transcription of messenger RNA from template DNA
  is catalyzed by RNA polymerase, which
• Separate the two strands of DNA and link RNA
  nucleotides as they base-pair along the template
• Add nucleotides to the 3 end thus mNRA grows in
  the 5 3 direction

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• Transcription occurs in three stages
• Polymerase binding and initiation
• Elongation
• Termination

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• In eukaryotes, RNA polymerase cannot recognize
  the promoter ( about 100 nucleotides long)
  without transcription factors.
• RNA polymerase II cannot recognize the promotor
  site without binding to the TATA box ( a short
  sequence of nucleotides rich in A and T that is
  about 25 nucleotides upstream from the initiation
  site.

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RNA SPLICING

• Introns noncoding segments that lie between the
  coding segments
• Exons -- coding segments that are expressed
• Roberts and Sharp independently found evidence
  for split gene and both received a Nobel Prize

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• RNA polymerase transcribes all the sequences and
  this is known as pre mRNA. This pre mRNA never
  leaves the nucleus.
• The introns are removed and the abridged
  version of mRNA moves to the cytoplasm as the
  primary transcription.
• There are signals for the RNA splicing at the
  ends of the introns. Particles called small
  nuclear ribonucleoproteins (snRNPs snurps)
  begin the process.

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• Several snRNA molecules join to form a
  spliceosome ( about the size of a ribosome).
  These cut the introns and connect the exons.

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Ribozymes

• RNA molecules that function as enzymes
• Serve as a catalyst
• Makes the following statement obsolete
• All biological catalyst are proteins.

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What is the function of Introns and Splicing?

• May control gene activity and may regulate
  passage of mRNA to cytoplasm
• May have a role in the evolution of new proteins
• Increases the probability of crossing-over
• Split genes have a higher frequency of
  recombination.

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Translation RNA directed synthesis of a
polypeptide

• tRNA serves as the interpreter
• The role of tRNA is to transfer amino acids from
  the pool in the cytoplasm to a ribosome.
• The ribosome adds each amino acid to the growning
  end of a polypeptide chain ( 3)

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Transfer RNA

• Molecules of tRNA are not all identical
• Each type of tRNA links a mRNA codon with a
  specific amino acid.
• One end has an attachment site for amino acid and
  the anticodon is on the other end.
• May be used repeatedly
• Consists of a single RNA strand about 80
  nucleotides long

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• Looks like a clover leaf
• Loop protruding from L end is the anticodon and
  this binds to the mRNA
• There are 45 tRNA molecules. ( Should be 61 if
  there was a tRNA for each codon)
• Some tRNA can recognize 2 or more codons.

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• WOBBLE Allows for the relaxation of the
  base-pairing rules
• U can pair with A or G in the third position
• The most versatile tRNA has the inosin (I). The
  I ( in the wobble position) can form hydrogen
  bonds with U, C or A

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Steps in Translation

1. Correct match between tRNA and a amino acid. The
   amino acid is bound to the tRNA by aminocyl tRAN
   synthetase. There are 20 of these one for each
   amino acid
2. The codon-anticodon bonding.

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Ribosomes

• 2 subunits
• Large subunit
• Small subunit
• 60 of weight of ribosome is rRNA
• 3 Binding sites for tRNA
• P holds the tRNA with the growing polypetide
  chain
• A hold the next amino acid to be added
• E exit site

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• A single ribosome can make an average sized
  polypeptide in less than one minute.
• Usually a single mRNA is used to make many copies
  of a polypeptide simultaneously because several
  ribosomes work on translating at the same time.
• Polyribosome string of ribosomes
• .

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MutationsChange in the genetic makeup

• Point mutation change in one or a few
  base-pairs in a single gene
• If occurs in a gamete then the change is passed
  to the future.
• If it has an adverse effect then it is known as a
  genetic disorder or hereditary disease

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Types of Point Mutations

• Substitution
• Misssense still codes for an amino acid
• Nonsense does not code for an amino acid but
  changes to a code for STOP. This makes the chain
  too short and usually leads to nonfunctioning
  proteins.

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• Insertion or Deletion
• Frameshift not in a multiple of three

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• Spontaneous mutation are the result of errors in
  the DNA replication or repair.
• Mutagens are chemical or physical agents that
  cause DNA to change.
• AMES test is a test for mutagenic activity of
  different chemicals.