AP Biology 2005-2006 Chapter 17. From Gene to Protein.
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Transcript of AP Biology 2005-2006 Chapter 17. From Gene to Protein.
2005-2006 AP Biology
Chapter 17.
From Gene to Protein
2005-2006AP Biology
Metabolism teaches us about genes Metabolic defects
studying metabolic diseases suggested that genes specified proteins
alkaptonuria (black urine from alkapton) PKU (phenylketonuria)
each disease is caused by non-functional enzyme
A B C D E
Genescreate
phenotype
2005-2006AP Biology
2005-2006AP Biology
1 gene – 1 enzyme hypothesis Beadle & Tatum
Compared mutants of bread mold, Neurospora fungus
created mutations by X-ray treatments X-rays break DNA inactivate a gene
wild type grows on “minimal” media sugars + required precursor nutrient to synthesize
essential amino acids mutants require added amino acids
each type of mutant lacks a certain enzyme needed to produce a certain amino acid
non-functional enzyme = broken gene
2005-2006AP Biology
Beadle & Tatum 1941 | 1958
George Beadle
Edward Tatum
2005-2006AP Biology
Beadle & Tatum’s Neurospora experiment
2005-2006AP Biology
So… What is a gene? One gene – one enzyme
but not all proteins are enzymes but all proteins are coded by genes
One gene – one protein but many proteins are composed of several
polypeptides but each polypeptide has its own gene
One gene – one polypeptide but many genes only code for RNA
One gene – one product but many genes code for
more than one product …
Where doesthat leave
us?!
2005-2006AP Biology
Defining a gene…“Defining a gene is problematic because… one gene can code for several protein products, some genes code only for RNA, two genes can overlap, and there are many other complications.”
– Elizabeth Pennisi, Science 2003
gene
polypeptide 1
polypeptide 2
polypeptide 3
RNAgeneIt’s hard to
hunt for wabbits,if you don’t knowwhat a wabbitlooks like.
2005-2006AP Biology
proteinRNA
The “Central Dogma”
DNAtranscription translation
replication
How do we move information from DNA to proteins?
For simplicity sake,let’s go back togenes that codefor proteins…
2005-2006AP Biology
From nucleus to cytoplasm… Where are the genes?
genes are on chromosomes in nucleus Where are proteins synthesized?
proteins made in cytoplasm by ribosomes How does the information get from
nucleus to cytoplasm? messenger RNA
nucleus
2005-2006AP Biology
RNA ribose sugar N-bases
uracil instead of thymine U : A C : G
single stranded mRNA, rRNA, tRNA,
siRNA….
RNADNAtranscription
2005-2006AP Biology
Transcription Transcribed DNA strand = template strand
untranscribed DNA strand = coding strand Synthesis of complementary RNA strand
transcription bubble Enzyme
RNA polymerase
2005-2006AP Biology
Role of promoter1. Where to start reading
= starting point
2. Which strand to read
= template strand
3. Direction on DNA
= always reads DNA 3'5'
Transcription in Prokaryotes Initiation
RNA polymerase binds to promoter sequence on DNA
2005-2006AP Biology
Transcription in Prokaryotes Promoter sequences
RNA polymerase molecules bound to bacterial DNA
2005-2006AP Biology
Transcription in Prokaryotes Elongation
RNA polymerase unwinds DNA ~20 base pairs at a time
reads DNA 3’5’ builds RNA 5’3’ (the energy governs the synthesis!)
No proofreading 1 error/105 bases many copies short life not worth it!
2005-2006AP Biology
Transcription
RNA
2005-2006AP Biology
Transcription in Prokaryotes Termination
RNA polymerase stops at termination sequence
mRNA leaves nucleus through pores
RNA GC hairpin turn
2005-2006 AP Biology
Transcription in Eukaryotes
2005-2006AP Biology
Prokaryote vs. Eukaryote genes Prokaryotes
DNA in cytoplasm circular
chromosome naked DNA
no introns
Eukaryotes DNA in nucleus linear
chromosomes DNA wound on
histone proteins introns vs. exons
eukaryoticDNA
exon = coding (expressed) sequence
intron = noncoding (inbetween) sequence
2005-2006AP Biology
Transcription in Eukaryotes 3 RNA polymerase enzymes
RNA polymerase I only transcribes rRNA genes
RNA polymerase I I transcribes genes into mRNA
RNA polymerase I I I only transcribes rRNA genes
each has a specific promoter sequence it recognizes
2005-2006AP Biology
Transcription in Eukaryotes Initiation complex
transcription factors bind to promoter region upstream of gene
proteins which bind to DNA & turn on or off transcription
TATA box binding site only then does RNA
polymerase bind to DNA
2005-2006AP Biology
A A A A A3' poly-A tail
CH3
mRNA
5'
5' cap
3'
G PPP
Post-transcriptional processing Primary transcript
eukaryotic mRNA needs work after transcription Protect mRNA
from RNase enzymes in cytoplasm add 5' cap add polyA tail
Edit out introns
eukaryoticDNA
exon = coding (expressed) sequence
intron = noncoding (inbetween) sequence
primary mRNAtranscript
mature mRNAtranscript
pre-mRNA
spliced mRNA
50-250 A’s
2005-2006AP Biology
Transcription to translation Differences between
prokaryotes & eukaryotes time & physical separation
between processes RNA processing
2005-2006AP Biology
Translation in Prokaryotes Transcription & translation are
simultaneous in bacteria DNA is in
cytoplasm no mRNA
editing needed
2005-2006AP Biology
mRNA
From gene to protein
DNAtranscription
nucleus cytoplasm
mRNA leaves nucleus through nuclear pores
proteins synthesized by ribosomes using instructions on mRNA
aa
aa
aa
aa
aa
aa
aa
aa
ribosome
proteintranslation
2005-2006AP Biology
How does mRNA code for proteins?
TACGCACATTTACGTACGCGGDNA
AUGCGUGUAAAUGCAUGCGCCmRNA
Met Arg Val Asn Ala Cys Alaprotein
?
How can you code for 20 amino acids with only 4 nucleotide bases (A,U,G,C)?
2005-2006AP Biology
Cracking the code Nirenberg & Matthaei
determined 1st codon–amino acid match UUU coded for phenylalanine
created artificial poly(U) mRNA added mRNA to test tube of
ribosomes, tRNA & amino acids mRNA synthesized single
amino acid polypeptide chain
1960 | 1968
phe–phe–phe–phe–phe–phe
2005-2006AP Biology Marshall NirenbergHeinrich Matthaei
2005-2006AP Biology
Translation Codons
blocks of 3 nucleotides decoded into the sequence of amino acids
2005-2006AP Biology
AUGCGUGUAAAUGCAUGCGCCmRNA
mRNA codes for proteins in triplets
TACGCACATTTACGTACGCGGDNA
AUGCGUGUAAAUGCAUGCGCCmRNA
Met Arg Val Asn Ala Cys Alaprotein
?
2005-2006AP Biology
The code For ALL life!
strongest support for a common origin for all life
Code is redundant several codons for
each amino acid
Why is this a good thing?
Start codon AUG methionine
Stop codons UGA, UAA, UAG
2005-2006AP Biology
How are the codons matched to amino acids?
TACGCACATTTACGTACGCGGDNA
AUGCGUGUAAAUGCAUGCGCCmRNA
aminoacid
tRNA
anti-codon
codon
5' 3'
3' 5'
3' 5'
UAC
MetGCA
ArgCAU
Val
2005-2006AP Biology
proteinaa
aa
aa
aa
aa
aa
aa
aaaa
aa
aa
transcription
cytoplasm
nucleus
translation
2005-2006AP Biology
tRNA structure “Clover leaf” structure
anticodon on “clover leaf” end amino acid attached on 3' end
2005-2006AP Biology
Loading tRNA
Aminoacyl tRNA synthetase enzyme which bonds
amino acid to tRNA endergonic reaction
ATP AMP energy stored in
tRNA-amino acid bond unstable so it can release amino acid
at ribosome
2005-2006AP Biology
Ribosomes Facilitate coupling of
tRNA anticodon to mRNA codon organelle or enzyme?
Structure ribosomal RNA (rRNA)
& proteins 2 subunits
large small
2005-2006AP Biology
Ribosomes P site (peptidyl-tRNA site)
holds tRNA carrying growing polypeptide chain
A site (aminoacyl-tRNA site) holds tRNA carrying next amino acid to
be added to chain E site (exit site)
empty tRNA leaves ribosome from exit site
2005-2006AP Biology
Building a polypeptide Initiation
brings together mRNA, ribosome subunits, proteins & initiator tRNA
Elongation Termination
2005-2006AP Biology
Elongation: growing a polypeptide
2005-2006AP Biology
Termination: release polypeptide Release factor
“release protein” bonds to A site bonds water molecule to polypeptide chain
Now what happens to the polypeptide?
2005-2006AP Biology
Protein targeting Signal peptide
address label
Destinations: secretion nucleus mitochondria chloroplasts cell membrane cytoplasm
start of a secretory pathway
2005-2006AP Biology
Can you tell the story?
DNA
pre-mRNA
ribosome
tRNA
aminoacids
polypeptide
mature mRNA
5' cap
polyA tail
large subunit
small subunit
aminoacyl tRNAsynthetase
E P A
5'
3'
RNA polymerase
exon intron
tRNA
2005-2006AP Biology
Put it all together…
2005-2006 AP Biology
Any Questions??
2005-2006 AP Biology
Chapter 17.
Mutations
2005-2006AP Biology
Code is redundant several codons for
each amino acid “wobble” in the tRNA “wobble” in the
aminoacyl-tRNA synthetase enzyme that loads the tRNA
Universal code
2005-2006AP Biology
Mutations
When do mutations affect the next generation?
Point mutations single base change base-pair
substitution silent mutation
no amino acid change redundancy in code
missense change amino acid
nonsense change to stop codon
2005-2006AP Biology
Point mutation leads to Sickle cell anemiaWhat kind of mutation?
2005-2006AP Biology
Sickle cell anemia
2005-2006AP Biology
Mutations Frameshift
shift in the reading frame
changes everything “downstream”
insertions adding base(s)
deletions losing base(s)
2005-2006 AP Biology
What’s thevalue of
mutations?
2005-2006 AP Biology
Chapter 17.
RNAProcessing
2005-2006AP Biology
Transcription -- another look The process of transcription includes
many points of control when to start reading DNA where to start reading DNA where to stop reading DNA editing the mRNA protecting mRNA as it travels through
cell
2005-2006AP Biology
Primary transcript Processing mRNA
protecting RNA from RNase in cytoplasm add 5’ cap add polyA tail
remove introns
AUG UGA
2005-2006AP Biology
Protecting RNA 5’ cap added
G trinucleoside (G-P-P-P) protects mRNA
from RNase (hydrolytic enzymes)
3’ poly-A tail added 50-250 A’s protects mRNA
from RNase (hydrolytic enzymes) helps export of RNA from nucleus
UTR UTR
2005-2006AP Biology
Dicing & splicing mRNA Pre-mRNA mRNA
edit out introns intervening sequences
splice together exons expressed sequences
In higher eukaryotes 90% or more of gene can be intron no one knows why…yet
there’s a Nobel prize waiting…
“AVERAGE”…“gene” = 8000bpre-mRNA = 8000bmature mRNA = 1200bprotein = 400aalotsa “JUNK”!
2005-2006AP Biology
Discovery of Split genes1977 | 1993
Richard Roberts Philip Sharp
NE BioLabs MITadenovirus
common cold
2005-2006AP Biology
snRNPs small nuclear RNA RNA + proteins
Spliceosome several snRNPs recognize splice site
sequence cut & paste
RNA as ribozyme some mRNA can
splice itself RNA as enzyme
Splicing enzymes
2005-2006AP Biology
Ribozyme RNA as enzyme
Sidney Altman Thomas Cech
1982 | 1989
Yale U of Colorado
2005-2006AP Biology
Splicing details No room for mistakes!
editing & splicing must be exactly accurate a single base added or lost throws off the
reading frame
AUG|CGG|UCC|GAU|AAG|GGC|CAU
AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGUCCGAUAAGGGCCAU
AUG|CGG|GUC|CGA|UAA|GGG|CCA|U
AUGCGGCTATGGGUCCGAUAAGGGCCAUAUGCGGGUCCGAUAAGGGCCAU
Met|Arg|Ser|Asp|Lys|Gly|His
Met|Arg|Val|Arg|STOP|
2005-2006AP Biology
Alternative splicing Alternative mRNAs produced from same gene
when is an intron not an intron… different segments treated as exons
Hardto definea gene!
2005-2006AP Biology
Domains Modular architecture
of many proteins separate
functional & structural regions
coded by different exons in same “gene”
2005-2006AP Biology AAAAAAAAGTP
20-30b
3'
promoter transcriptionstop
transcriptionstart
introns
The Transcriptional unit (gene?)
transcriptional unitTAC ACT
DNA
DNATATA5'RNA
polymerase
pre-mRNA
5' 3'
translationstart
translationstop
mature mRNA
5' 3'
UTR UTR
exonsenhancer
1000+b
2005-2006 AP Biology
Any Questions??
2005-2006AP Biology
20-30b
3'
introns
The Transcriptional unit
transcriptional unitTAC ACTDNATATA
5'RNApolymerase
5' 3'
5' 3'
exonsenhancer
1000+b