Protein Synthesis DNA RNA Protein

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Protein SynthesisDNA ? RNA ? Protein

• Chapter 17

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• The info portion of DNA is in the specific
  sequences of nucleotides along DNA strands
• Proteins are the links between genotype and
  phenotype
• Beadle and Tatum established link between genes
  and enzymes

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Beadle and Tatum

• They mutated Neurospora crassa, a bread mold,
  with X-rays and screened the survivors for
  mutants that differed in their nutritional needs
• Wild-type Neurospora can grow on a minimal medium
  of agar, inorganic salts, glucose, and the
  vitamin biotin

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Beadle and Tatum

• Mutants that cannot synthesize their own argenine
  (amino acid) could if certain intermediates were
  placed in the MM agar
• This not only showed that metabolic processes
  were step-wise, but also that the synthesis of
  aas, and therefore proteins, were a result of
  the organisms genetic information the one gene
  one enzyme hypothesis

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Beadle and Tatum
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Beadle and Tatum

• Since not all proteins are enzymes, more research
  refined the hypothesis to the one gene one
  protein
• Even more research has shown that most proteins
  are several polypeptides (4O structure) working
  together
• Therefore, the hypothesis is now the one gene
  one polypeptide hypothesis

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Protein Synthesis

• Genes code for polypeptides
• RNA is the link between the DNA (in the nucleus)
  and the ribosomes (in the cytoplasm)

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Protein Synthesis

• RNA has three basic differences
  from DNA

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Protein Synthesis

• To get from DNA (written in one chemical
  language) to protein (written in
  another) requires two major
  stages
• Transcription
• Translation

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Transcription

• Process by which a molecule of RNA is synthesized
  that is complementary to a specific sequence of
  DNA
• Involves 3 stages initiation, elongation
  termination
• Used to synthesize any type of RNA from a DNA
  template
• Transcription of a gene produces a messenger RNA
  (mRNA) molecule

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Transcription Initiation

• RNA polymerase attaches to a binding site in a
  promoter upstream from the gene on DNA strand
• The presence of the promoter determines which
  strand of the DNA acts as the template

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Transcription Initiation

• In eukaryotes, transcription factors (proteins)
  recognize and bind to the promotor, especially a
  TATA box.
• RNA polymerase then binds to
  transcription factors to create
  atranscriptioninitiation complex

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Transcription Initiation

• In prokaryotes, RNA polymerase can recognize and
  bind directly to the promotor region
• RNA polymerase then starts transcription in both
  types of cells

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Transcription Initiation

• A short section of DNA is unwound
• Free RNA nucleotides move in and H-bond to
  complementary bases on the DNA template strand

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Transcription Elongation

• RNA polymerase links RNA nucleotides together in
  a 5 to 3 direction (reads the DNA
  the gene 3 to 5, just like
  DNA polymerase)
• The enzyme unzips 10-20 base pairs
  at a time

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Transcription Elongation

• Behind the site of RNA synthesis, the double
  helix re-forms and the RNA molecule peels away

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Transcription Termination

• RNA polymerase detaches when it reaches a
  terminator
• Completed RNA molecule is released from DNA
  template

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Transcription

• A single gene can be transcribed by several RNA
  polymerases simultaneously more RNA produced in
  less time

• This slide also shows several ribosomes
  performing translation simultaneously on the
  newly formed mRNA molecules

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Transcription

• A single gene can be transcribed by several RNA
  polymerases simultaneously more RNA produced in
  less time
• The length of each new strand reflects how much
  of the gene has been transcribed

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Transcription
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Transcription

• Occurs in the nucleus of eukaryotic and cytoplasm
  of prokaryotic cells
• Is regulated by operons (bacterial
  cells) or transcription factors
  (multicellular organisms)
• The basic mechanics of transcription and
  translation are similar in eukaryotes and
  prokaryotes

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Transcription

• Bacteria have a single type of RNA polymerase
  that synthesizes all RNA molecules
• In contrast, eukaryotes have three RNA
  polymerases (I, II, and III) in their nuclei
• RNA polymerase II is used for mRNA synthesis

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Transcription modification

• Before the RNA is sent out to the cytoplasm to be
  translated, it is modified
• At the 5 end of the mRNA, a modified form of
  guanine is added the 5 cap
• Protects mRNA from hydrolytic enzymes
• Also functions as an attach here signal for
  ribosomes

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Transcription modification

• At the 3 end, an enzyme adds 50 to 250 adenine
  nucleotides the poly-A tail
• Also inhibits hydrolysis
• The mRNA molecule also includes nontranslated
  leader and trailer segments

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Transcription modification

• RNA splicing excises noncoding introns from the
  coding exons

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Transcription modification

• Spliceosomes do RNA splicing
• consist of a variety of proteins and several
  small nuclear ribonucleoproteins (snRNPs)
• Each snRNP has several protein molecules and a
  small nuclear RNA molecule (snRNA)
• Each is about 150 nucleotides long

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Transcription modification

• Pre-mRNA combineswith snRNPs and otherproteins
  to form aspliceosome
• Within the spliceosome, snRNA
  base-pairs withnucleotides at the ends
  of the intron
• The RNA transcript is cut to release the intron,
  and the exons are spliced together
  the spliceosome then comes
  apart, releasing mRNA, which now
  contains only exons

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Transcription modification

• Alternative RNA splicing gives rise to two or
  more different polypeptides, depending on which
  segments are treated as exons
• Early results of the Human Genome Project
  indicate that this phenomenon may be common in
  humans

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RNA

• Three major types are transcribed mRNA, rRNA,
  and tRNA
• mRNA (messenger RNA) encodes genetic info from
  DNA and carries it into the cytoplasm
• Each three consecutive mRNA bases forms a genetic
  code word (codon) that codes for a particular
  amino acid

codon
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RNA

• rRNA (ribosomal RNA) associates with proteins
  to form ribosomes
• Subunits are separate in the cytoplasm, but join
  during protein synthesis (translation)

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RNA

• tRNA (transfer RNA) transports specific amino
  acids to ribosome during protein synthesis
  (translation)
• Anticodon - specific sequence of 3 nucleotides
  complementary to codon determines the specific
  amino acid that binds to tRNA

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Types of RNA
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Translation

• Process by which an mRNA sequence is translated
  into an amino acid sequence (polypeptide/protein)
• Occurs in the cytoplasm of eukaryotic and
  prokaryotic cells
• Requires mRNA, tRNAs, amino acids, and ribosomes
• Involves 3 stages
• Initiation
• Elongation
• Termination

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Translation Initiation

• Small subunit attaches to initiator site (codon)
  on mRNA AUG
• Large subunit attaches to complex and attracts
  the initiator tRNA carries methionine

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• Each ribosome has a binding site for mRNA and
  three binding sites for tRNA molecules.
• The P site holds the tRNA carrying the growing
  polypeptide chain.
• The A site carries the tRNA with the next amino
  acid.
• Discharged tRNAs leave the ribosome at the E site.

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Translation Elongation

• Elongation has three repetitive parts
• Codon recognition
• Peptide bond formation
• Translo- cation

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Translation Elongation

• During codon recognition, an elongation factor
  assists H-bonding between the mRNA codon under
  the A site with the corresponding anticodon of
  tRNA carrying the appropriateamino acid
• This step requires the hydrolysis of two GTP

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Translation Elongation

• 61 of 64 triplets code for amino acids
• The codon AUG not only codes for the amino acid
  met, but also indicates
  the start of translation
• Three codons do not indicate amino acids but
  signal the termination of translation.

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Translation Elongation

• During peptide bond formation, an rRNA molecule
  catalyzes the formation of a peptide bond between
  the polypeptide in the P site with the new amino
  acid in the A site
• This step separates the tRNA at the P site from
  the growing polypeptide chain and transfers the
  chain, now one amino acid longer, to the tRNA at
  the A site

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Translation Elongation

• During translocation, the ribosome moves the tRNA
  with the attached polypeptide from the A site to
  the P site
• Because the anticodon remains bonded to the mRNA
  codon, the mRNA moves along with it
• The next codon is now available at the A site
• The tRNA that had been in the P site is moved to
  the E site and then leaves the ribosome
• Translocation is fueled by the hydrolysis of GTP

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Translation Termination

• Termination occurs when one of the three stop
  codons reaches the A site.
• A release factor binds to the stop codon and
  hydrolyzes the bond between the polypeptide and
  its tRNA in the P site.
• This frees the polypeptide and the translation
  complex disassembles

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Translation

• Usually, several copies of the polypeptide/protein
  are made at a time.

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Translation

• Because bacteria lack nuclei, transcription and
  translation are coupled
• Ribosomes attach to the leading end of a mRNA
  molecule while transcription is still in progress

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Review
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Review

• The genetic code is redundant but not ambiguous
• Several different codons can code for the same
  amino acid
• However, any one codon codes for only one
  specific amino acid
• I.e., If you have a specific codon, you can be
  sure of the corresponding amino acid if you know
  only the amino acid, there may be several
  possible codons

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Mutation

• Any change in the original genetic code (sequence
  of nucleotides) May not affect phenotype (silent
  mutation).
• Can affect somatic cells (somatic mutation) or
  sex cells (germinal mutation)
• Mutations in gametes are only kinds that can be
  passed to offspring
• Can form spontaneously or be induced by a mutagen

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Mutation

• Point mutation mutation at one base pair
• Base pair substitution one set of complements
  is replaced with the other
• Silent mutation change in DNA does not change
  the amino acid it codes for
• Missense
• Nonsense

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Mutation

• Point mutation mutation at one base pair
• Missense mutations changes a codon to specify a
  different amino acid
• Ex sickle cell anemia

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Mutation

• Nonsense mutation change an amino acid codon
  into a stop codon, nearly always leading to a
  nonfunctional protein

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Mutation

• Insertions or deletions adding or subtracting
  nucleotides
• Frameshift mutation the insertion or deletion
  of DNA nucleotides results in disruption of the
  reading frame
• Ex. cystic fibrosis

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