Protein Synthesis (From Nucleus to Cytoplasm)

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Protein Synthesis(From Nucleus to Cytoplasm)
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The Central Dogma

• www.video.sina.com.cn

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Amino Acids
tRNA
TRANSCRIPTION
TRANSLATION
DNA
mRNA
PROTEIN
Ribosomes
REPLICATION
Protein
rRNA
The Central Dogma (in equation form)
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• Initial Problem
• DNA codes for proteins BUT DNA is confined to
  the nucleus and the equipment to carry out
  protein synthesis is in the cytoplasm.
• (The equipment includes ribosomes, tRNA, and
  amino acids).
• Solution???
• DNA can be copied into an RNA molecule
  (messenger RNA (mRNA)), which can travel from the
  nucleus to the cytoplasm carrying the
  instructions for building a protein.
• This DNA to mRNA copying process is known as
  transcription (see fig. 25.7 on p. 512).

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Visual Examples of Transcription
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Transcription

• During this process, DNA serves as a template
  (guide) for the production of mRNA
• The enzyme DNA helicase serves to unwind and
  unzip the portion of the DNA double helix (ie.
  The GENE) that is to be transcribed (copied into
  mRNA).
• Once this occurs, free-floating RNA nucleotides
  (within the nucleus along with free-floating DNA
  nucleotides) bond to the exposed bases of DNA
  through complementary base-pairing (forming
  hydrogen bonds).
• RNA nucleotides bind to only ONE exposed DNA
  strand.

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• A (mRNA) binds with T (DNA).
• U (mRNA) binds with A (DNA).
• C (mRNA) binds with G (DNA).
• G (mRNA) binds with C (DNA).

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• The first three mRNA bases are always AUG (called
  the start codon), which means that the first
  three DNA bases that are transcribed are TAC.
• The last three mRNA bases are always one of UAG,
  UAA, or UGA (STOP codons), which means that the
  last three DNA bases transcribed are always one
  of ATC, ATT, or ACT.
• The enzyme RNA Polymerase then works to join
  added mRNA nucleotides to each other
  (sugar-P-sugar-P etc) through dehydration
  synthesis (producing water).
• So, the mRNA nucleotides form spontaneous H-bonds
  with exposed DNA bases but then need enzymatic
  aid (RNA polymerase) to form the actual mRNA
  chain.

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• DNA helicase continues to unwind/unzip the DNA
  until the gene that requires copying has been
  fully exposed.
• The signal for DNA helicase to stop is when it
  encounters ATC, ATT, or ACT.
• A poly-adenine tail is added to one end of the
  finished mRNA while a guanine-based cap is
  added to the other side to protect the molecule
  from cytoplasmic enzymes.
• Once this is finished, the mRNA moves into the
  cytoplasm, through a nuclear pore, and the DNA
  joins back together by reforming its
  complementary base-pairing H-bonds.

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START Codon
STOP Codon
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The Genetic Code

• Query How is a sequence of nitrogenous bases on
  mRNA going to be used to code for a sequence of
  amino acids and hence, a protein???
• First of all, DNA is the universal code (ie.
  Every living thing has DNA, and DNA codes for
  proteins (through the use of mRNA)).
• There are 20 amino acids in nature.
• There are 4 different nitrogenous bases in both
  DNA and mRNA, and they serve as a code for the
  amino acid sequence of proteins.

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• The code reads as a TRIPLET CODE (43 64
  possibilities), meaning that three nitrogenous
  bases, as a group, code for ONE amino acid.
• A singlet code could only code for 41 4 amino
  acids (yet, there are 20) thus proving
  inadequate.
• A doublet code could only code for 42 16 amino
  acids.
• A group of three mRNA nucleotides (bases) is
  called a CODON. In total, there are 64 (43)
  different codons.
• A group of three DNA nucleotides (bases) is
  called a DNA triplet.

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• There are 61 different codons that correspond to
  the 20 different amino acids (one of these 61 is
  AUG, which is the start codon, that codes for
  the amino acid methionine).
• The other three codons are called STOP codons,
  which terminate the formation of the
  polypeptide/protein chain.
• These STOP codons DO NOT code for an amino acid.
  
• The Genetic code is sometimes referred to as
  being redundant, because most amino acids are
  coded for by 2-6 different codons.

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• AAA GCU ACC GGU UAC GUC UAG mRNA sequence
• Questions i. How many codons present?
  ii. How many a. acids coded for?
• iii. What is the a. acid sequence?
• lysine, alanine, threonine, glycine,
• tyrosine, valine, STOP

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• iv. What was the DNA sequence of triplets that
  coded for this mRNA?
• TTT CGA TGG CCA ATG CAG ATC

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Translation

• The process of turning mRNA into a protein (ie.
  Translating the language of nitrogenous bases
  into the language of amino acids).
• Recall that mRNA is constructed in the nucleus
  through the process of transcription, and is sent
  out of the nucleus through a nuclear pore.
• Once mRNA enters the cytoplasm, it immediately
  associates with a ribosome (either a free
  ribosome or a Rough ER ribosome).
• The ribosome attaches to the mRNA at the
  guanine-based cap that not only served as
  protection from enzymes, but acted as a start
  here signal.

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• Translation requires two other types of RNA
• rRNA (Ribosomal RNA)
• -- joins with ribosomal proteins (from
  nucleolus) to form ribosomes.
• -- produced in the nucleolus.
• -- one ribosome has two subunits
• a. Large subunit (3 rRNAs and proteins)
• b. Small subunit (1 rRNA and proteins)
• -- the two subunits remain close together but do
  not actually attach until just prior to
  translation.
• -- rRNA is not involved in any coding or
  translating, it is purely structural.

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• ii. tRNA (Transfer RNA)
• -- located in the cytoplasm and serve as
  carriers of singular amino acids to the
  mRNA/ribosome complex.
• -- tRNAs carry one amino acid at one end, and a
  specific ANTICODON at the other end.
• -- this anticodon will match-up with a
  complementary codon on mRNA (through
  complementary base-pairing) (fig. 25.8 p. 513).
• -- tRNA molecules are very specific (ie. A
  tRNA with a certain anticodon will ALWAYS be
  carrying the same amino acid).
• -- remember, though, that the translation table
  is translated from mRNA codons, not the tRNA
  anticodons.

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• Eg. If the mRNA codon being translated is ACG,
  what anticodon and amino acid will the tRNA
  molecule, specific to this codon, be carrying?
• Ans. Anticodon UGC
• Amino Acid Threonine (need table)
• Eg 2. Codon CAA, find anticodon and a. acid.
• Anticodon GUU, A. acid Glutamine
• Eg 3. DNA triplet TTC, find mRNA codon, tRNA
  anticodon, and amino acid.
• mRNA codon AAG, anticodon UUC
• Amino acid Lysine

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• HINT The anticodon will be the same as the
  original DNA triplet except that a U will replace
  a T.

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Three Steps of Translation

• INITIATION
• -- the cap of mRNA binds to the ribosome and
  the ribosome moves along the mRNA, reading it,
  until it comes upon the start codon, AUG.
• -- the tRNA with anticodon UAC binds to the AUG
  codon (through complementary base-pairing) at the
  A-site, and delivers the first amino acid
  Methionine.
• -- see handout (crude handout, that is)

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A-site
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• ELONGATION (lengthening of the amino acid chain).
• -- firstly, the ribosome is large enough to
  accommodate two tRNAs at the same time ? the
  incoming tRNA (at the A-site) and the
  outgoing tRNA (at the P-site).
• -- after the first tRNA binds to the mRNA codon,
  the ribosome shifts one codon (3 bases), thus
  exposing a new codon in the A-site which can then
  be bonded to by a new tRNA with the complementary
  anticodon.
• -- after this next tRNA binds, the ribosome
  shifts again and bumps the tRNA in the P-site
  off of the ribosome.

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• -- before it is bumped off of the ribosomes
  P-site, the outgoing tRNA molecule always passes
  the amino acid chain, via dehydration synthesis,
  to the tRNA that is shifting from the A-site to
  the P-site.
• -- the liberated or bumped tRNA (now without an
  amino acid) will eventually pick up the same
  amino acid that it just ceded and will rejoin the
  group of tRNAs waiting to be chosen.
• -- amino acids are readily available in the
  cytoplasm.
• -- the ribosome continually shifts to accept more
  tRNA molecules so that the protein chain can grow
  one amino acid at a time.

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P-site
A-site
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• iii. TERMINATION
• -- occurs once the codon appearing in the A-site
  of the ribosome is a STOP codon.
• -- this STOP codon is recognized by the ribosome
  complex and a RELEASE FACTOR protein is summoned
  from the cytoplasm into the A-site.
• -- there is NO tRNA molecule for these codons.
• -- once the release factor protein binds to the
  STOP codon, the ribosome dissociates into its two
  subunits and falls off the mRNA (which is
  recycled).
• -- the peptide/protein chain is released by the
  tRNA in the P-site into the lumen of the Rough ER
  (if for export), or into a transition vesicle
  bound for the Golgi for modifications (if it is
  to remain in the cell).
• -- see fig. 25.9 p. 514, fig 25.11 p. 516
• -- read translation summary on pp. 512-516.

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By simply eating protein, you can build new,
custom proteins and maybe look like this
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