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RNA and Protein Synthesis
How does DNA “code”?
DNA inherited by an organismLeads to specific traits by dictating
synthesis of proteins DNA directs protein synthesis/gene
expression2 stages - transcription and translation
Central Dogma
DNA RNA PROTEIN
History of RNA Synthesis George Beadle & Edward Tatum in
(‘40s)- ‘One gene, one enzyme’ Function of a gene is to dictate the
production of a specific enzyme
Beadle
1903-1989
American
Nobel Prize 1958
Tatum
1909-1975
American
Nobel Prize 1958
After Beadle & Tatum Some genes encode proteins that
are not enzymes One gene is responsible for one
polypeptide chain, and some proteins have more than one chain
One gene, one polypeptide hypothesis
The BIG Picture DNA is transcribed to form RNA RNA is translated to form protein
Prokaryotic cell - no nucleus, mRNA produced by transcription immediately translated without additional processing
TRANSLATION
TRANSCRIPTION DNA
mRNA
Ribosome
Polypeptide
Eukaryotic cell - nucleus provides a separate compartment for transcription. Original RNAtranscript (pre-mRNA) processed in various ways before leaving nucleus as mRNA
TRANSCRIPTION
RNA PROCESSING
TRANSLATION
mRNA
DNA
Pre-mRNA
Polypeptide
Ribosome
Nuclearenvelope
RNA RNA is used as an intermediary
between DNA and proteins RNA is a single strand nucleotide
polymer Composition
Sugar- RibosePhosphate group(s)Uracil substitutes for thymine
uracil
Transcription Overview A copy of the DNA is
made in the form of mRNA (messenger RNA) in transcription
Translation Overview Translation involves mRNA, tRNA
(transfer RNA), and rRNA (ribosomal RNA) coordinating to produce proteins
Translation Overview 2 mRNA has sequences of 3
nucleotides called codons Codons are read in sequences of 3
called triplet code
Translation Overview 3
Codons are written 5’ to 3’ fashion
Each codon codes for one amino acid
Codons do not overlap
Translation Overview 4 Four bases can combine in 43
combinations– more than enough to code for the 20 naturally occurring amino acids
43 = 64 Why don’t we have 64 amino acids?
Universal Code Genetic code (AA
code) nearly universalFrom simplest bacteria
to complex animals Genes can be
transcribed & translated after being transplanted from one species to another
Tobacco plant with firefly gene
Translation Overview - tRNA tRNA molecule has a
sequence of 3 nucleotides- the anticodon
Anticodons base pair with the codon in a complementary way
Anticodons are written in 3’ to 5’ direction
Translation Overview 6 Ribosomes are composed of proteins
and rRNA
The E, P, and A are rRNA
Ribosome
Transcription: Initiation
Synthesis of RNA from the DNA template
Main enzyme is RNA polymerase Transcription does not involve a
primer - it begins at a promoter site The promoter is a “start” sequence
Transcription: Elongation RNA synthesis proceeds in a 5’-3’
direction copying DNA from the 3’-5’
Upstream- towards 5’ end of mRNA sequence
Downstream- towards the 3’ end of mRNA sequence
Transcription: Elongation
RNA polymerase moves along the DNA
Untwists the double helix, exposing about 10 to 20 DNA bases at a time for pairing with RNA nucleotides
Transcription: Elongation Bacterial promoters are about 40
bases long and are located in the DNA just upstream from the starting point
Transcription: Termination Sequences at the end of the gene act as
stop signals Typically only one strand of DNA is
transcribed and is called the template strand
Termination (eukaryotes) Enzymes in nucleus modify pre-mRNA
(before genetic messages sent to the cytoplasm)
mRNA contains additional base sequences that do not directly code for proteins5’ end: modified nucleotide cap3’ end: poly-A tail
`
5’ Cap & Poly A Tail
5’ CapProtect mRNA from hydrolytic enzymesFunctions as an “attach here” signal for
ribosomes Poly A Tail
50 – 250 nucleotidesSame function as 5’ capFaciliates export from nucleus
RNA Modification
RNA splicingRemoves introns and joins exonsIntrons – noncoding regions of DNAExons – coding portions of DNA Introns stay “in” the nucleus, exons
“exit” the nucleus
Spliceosome
Splicing accomplished by a spliceosomeConsists 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
Translation 1 During translation, the nucleic acid
message is decoded An amino acid is attached to tRNA
before becoming incorporated into a polypeptideTo form a polypeptide chain, the amino
and carboxyl groups of amino acids are joined
Translation 2
The specific sequence of the amino acids (primary structure) is dictated by the sequence of codons of the mRNA
Translation 3
tRNA is linked to amino acids by aminoacyl-tRNA synthetases
This is an energy requiring process
RNA molecules - tRNA
RNA molecules have specialized regions with specific functions
tRNA molecules have attachment sites for amino acids
RNA molecules
tRNA molecules have anticodons that bind to complementary codons of the mRNA
If the mRNA codon is UAC, then what is the anticodon present on the tRNA?
RNA molecules tRNA must be recognized by both the
specific aminoacyl-tRNA synthetase and the ribosome
RNA molecules tRNA ~ 70 nucleotides
long, some generic sections & some unique sections
The nucleotide chain is folded back upon itself to form 3 or more loops with unpaired nucleotides exposed
Ribosomes Components of
translational machinery come together at the ribosomes
Ribosomes are composed of two subunits
Ribosomes The large subunit has a
groove into which the small subunit fits
Ribosomes are transcribed from DNA, but do not carry information
Function as physical site of translation & as a catalyst
Ribosome The A site of the
ribosome is where the aminoacyl-tRNA binds
The P site is where the tRNA holding the polypeptide chain is positioned
Translation Steps Inititiation Elongation Termination
Initiation Initiation factors
(proteins) move an initiation tRNA onto the small ribosomal subunit
The codon for the initiation is AUG, which codes for the amino acid methionine
Initiation 2 Initiation
complex binds to ribosome recognition sequences on the mRNA, and aligns anticodon of tRNA with the codon of mRNA
Initiation 3 The large
ribosomal subunit then binds, forming the functional ribosome
Elongation 1 Addition of new
amino acids Initiator tRNA is
bound to P site of the ribosome - A site is unoccupied until the next aminoacyl-tRNA moves in
Elongation 2 Energy for this
process comes from GTP
Peptide bond formation
Amino group of the new amino acid & carboxyl group of “old” amino acid
Elongation 3 Protein synthesis proceeds from the
amino end to the carboxyl end The tRNA molecule is released from
the P site requiring ribozyme, peptidyl transferase
Translocation - movement of the growing polypeptide chain from the A site to P site - energy comes from GTP
Elongation 4 Translation of the mRNA proceeds in
a 3’ to a 5’ direction, which is the same as the direction of transcription
Translocation ensures
Termination Occurs when the mRNA presents the
codons UAA, UGA, or UAG; No complementary tRNA Release factors recognize codons The ribosome dissociates into the
two subunits
Translation Tutorial http://telstar.ote.cmu.edu/Hughes/Hu
ghesArchive/tutorial/polypeptide/tutorial.swf
Polyribosome A single mRNA can make many copies
of a polypeptide simultaneously Multiple ribosomes, polyribosomes,
may trail along the same mRNA
Mutations Point mutations – single bp change Inheritable (if occurs in gametes)
Mutations are changes in DNA
Point mutation
Base pair substitutionReplacement of a pair of
complementary nucleotides with another nucleotide pair
Some have little/no impact Silent mutation AA change
SimilarNonessential
Missense mutation
AA change
Nonsense mutation
AA change to stop codon
Mutations Frameshift mutations- involve the
insertion or deletion of a base They result in an entirely different
sequence of amino acids because they change the _______________
Mutations are changes in DNA
Frameshift Mutations
Mutations Transposons - movable sequences
of DNA that may move into another area of DNAMay disrupt genes, but may inactivate
othersTransposons have some similarities to
retroviruses
Mutations are changes in DNA
Transposons
Barbara McClintock (1902-1992)
Nobel Prize 1983 Cornell University Discovered the
transposons “jumping genes”
Studied maize
Mutations
Hot spots - regions of DNA more likely to undergo mutations, and are often regions of repeated nucleotides, causing the polymerases to slip
Mutagens Agents that cause mutations
Including ionizing radiationMutations in somatic cells are not
passed on to the next generationSome mutagens are also carcinogens