View
221
Download
0
Category
Preview:
Citation preview
AP Biology 2007-2008
From Gene to Protein
How Genes Work
SLIDE SHOW BY KIM FOGLIA (modified)All Blue edged slides are Kim’s
(hyperlinks may have been added)
AP Biology
What do genes code for?
proteins cells bodies
How does DNA code for cells & bodies? how are cells and bodies made from the
instructions in DNA
DNA
AP Biology
The “Central Dogma” Flow of genetic information in a cell
How do we move information from DNA to proteins?
_____________
____________
_______________
proteinRNADNA trait
DNA gets all the glory,
but proteins do all the work!
AP Biology
Inheritance of metabolic diseases suggested that genes coded for enzymes each disease (phenotype) is caused by
non-functional gene product lack of an enzyme Tay sachs PKU (phenylketonuria) albinism
Am I just the sum of my proteins?
Metabolism taught us about genes
A B C D E
disease disease disease disease
enzyme 1 enzyme 2 enzyme 3 enzyme 4
metabolic pathway
AP Biology
Beadle & Tatum 1941 | 1958
George Beadle
Edward Tatum
"for their discovery that genes act by regulating definite chemical events"
________________________hypothesis
AP Biology
Wild-typeNeurospora
Minimalmedium
Select one ofthe spores
Grow oncomplete medium
Minimalcontrol
Nucleicacid
CholinePyridoxine Riboflavin Arginine
Minimal media supplemented only with…
ThiamineFolicacid
NiacinInositolp-Aminobenzoic acid
Test on minimalmedium to confirmpresence of mutation
Growth oncompletemedium
X rays or ultraviolet light
asexualspores
spores
Beadle & Tatum
create mutations
positive control
negative control
experimentalsmutatio
n identifi
ed
amino acidsupplements
AP Biology
mRNA
From gene to protein
DNAtranscription
nucleus cytoplasm
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
proteintranslation
ribosome
trait
AP Biology
RNA ____________ sugar N-bases
_______ instead of thymine U : A C : G
single stranded lots of RNAs
mRNA, tRNA, rRNA, siRNA…
RNADNAtranscription
3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
RIBOSOMAL RNA (rRNA)Made in ______________2 subunits (large & small)Combine with proteins to
form ribosomesBacterial ribosomes different
size than eukaryotic ribosomes Evidence for ENDOSYMBIOTIC THEORY Medically significant-some antibiotics target
bacterial ribosomes w/o harming host
rRNA and t-RNA images from Image from: Biology; Miller and Levine; Pearson Education publishing as Prentice Hall; 2006mRNA image from http://wps.prenhall.com/wps/media/tmp/labeling/1140654_dyn.gif
3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
TRANSFER RNA (tRNA)
________________ sequence matches CODON on mRNA to add correctamino acids during protein synthesis
AMINOACYL-tRNA SYNTHETASE Enzyme attaches a specific amino acid using energy from ATP
http://www-math.mit.edu/~lippert/18.417/lectures/01_Intro/
3 KINDS OF RNA HELP WITH INFO TRANSFER FOR PROTEIN SYNTHESIS
MESSENGER RNA (mRNA)carries code from DNA to ribosomes
AP Biology
Transcription Making mRNA
transcribed DNA strand = _____________ strand untranscribed DNA strand = coding strand
same sequence as RNA synthesis of complementary RNA strand
transcription bubble enzyme
RNA polymerase
template strand
rewinding
mRNA RNA polymerase
unwinding
coding strand
DNAC C
C
C
C
C
C
C
C CC
G
GG
G
G G
G G
G
G
GAA
AA A
A
A
A
A
A A
A
AT
T T
T
T
T
T
T
T T
T
T
U U
5
35
3
3
5build RNA 53
•Transcription
AP Biology
RNA polymerases 3 RNA polymerase enzymes
RNA polymerase 1 only transcribes rRNA genes makes ribosomes
RNA polymerase 2 transcribes genes into mRNA
RNA polymerase 3 only transcribes tRNA genes
each has a specific promoter sequence it recognizes
AP Biology
Which gene is read? Promoter region
binding site before beginning of gene TATA box binding site binding site for RNA polymerase
& transcription factors
Enhancer region binding site far
upstream of gene turns transcription
on HIGH
AP Biology
Transcription Factors Initiation complex
transcription factors bind to _____________region suite of proteins which bind to DNA hormones? turn on or off transcription
trigger the binding of RNA polymerase to DNA
•Transcription
AP Biology
Matching bases of DNA & RNA Match RNA bases to DNA
bases on one of the DNA strands
U
A G GGGGGT T A C A C T T T T TC C C CA A
U
UU
U
U
G
G
A
A
A C CRNA
polymerase
C
C
C
C
C
G
G
G
G
A
A
A
AA
5' 3'
AP Biology
Eukaryotic genes have junk! Eukaryotic genes are not continuous
exons = the real gene expressed / coding DNA
introns = the junk inbetween sequence
eukaryotic DNA
exon = coding (expressed) sequence
intron = noncoding (inbetween) sequence
intronscome out!
AP Biology
mRNA splicing
eukaryotic DNA
exon = coding (expressed) sequence
intron = noncoding (inbetween) sequence
primary mRNAtranscript
mature mRNAtranscript
pre-mRNA
spliced mRNA
Post-transcriptional processing eukaryotic mRNA needs work after transcription primary transcript = ______________ mRNA splicing
edit out _____________ make mature mRNA transcript
~10,000 bases
~1,000 bases
AP Biology
1977 | 1993
Richard Roberts Philip
SharpCSHL
MITadenovirus
common cold
Discovery of exons/introns
beta-thalassemia
AP Biology
Splicing must be accurate No room for mistakes!
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|
AP Biology
RNA splicing enzymes
snRNPs
exonexon intron
snRNA
5' 3'
spliceosome
exonexcisedintron
5'
5'
3'
3'
3'
lariat
exonmature mRNA
5'
No, not smurfs!“snurps”
______________ small nuclear RNA proteins
______________ several snRNPs recognize splice
site sequence cut & paste gene
Whoa! I think we just broke
a biological “rule”!
mRNA EDITING
ALL ENZYMES ARE PROTEINS?
______________-RNA molecules that function as enzymes (In some organisms pre-RNA can remove its own introns)
PROCESSING RNA
SPLICEOSOMES
AP Biology
Alternative splicing Alternative mRNAs produced from same gene
when is an intron not an intron… different segments treated as exons
Starting to gethard to
define a gene!
AP Biology
A A AA
A3' poly-A tail
mRNA
5'5' cap
3'
G PPP
50-250 A’s
More post-transcriptional processing Need to protect mRNA on its trip from
nucleus to cytoplasm enzymes in cytoplasm attack mRNA
protect the ends of the molecule add ____________ cap add ____________ tail
longer tail, mRNA lasts longer: produces more protein
AP Biology
mRNA
From gene to protein
DNAtranscription
nucleus cytoplasm
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
ribosome
trait
proteintranslation
AP 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)?
4
4
20
ATCG
AUCG
AP Biology
AUGCGUGUAAAUGCAUGCGCCmRNA
mRNA codes for proteins in triplets
TACGCACATTTACGTACGCGGDNA
AUGCGUGUAAAUGCAUGCGCCmRNA
Met Arg Val Asn Ala Cys Alaprotein
?
codon
AP Biology
Cracking the code1960 | 1968
Crick determined 3-letter (triplet) codon system
Nirenberg & Khorana
WHYDIDTHEREDBATEATTHEFATRATWHYDIDTHEREDBATEATTHEFATRAT
Nirenberg (47) & Khorana (17) determined mRNA–amino acid match added fabricated mRNA to test tube of
ribosomes, tRNA & amino acids created artificial UUUUU… mRNA found that UUU coded for phenylalanine
AP Biology
The code Code for ALL life!
strongest support for a common origin for all life
Code is redundant several codons for
each amino acid 3rd base “wobble”
Start codon AUG methionine
Stop codons UGA, UAA, UAG
Why is thewobble good?
AP 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
AP Biology
mRNA
From gene to protein
DNAtranscription
nucleus cytoplasm
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
aa
ribosome
traitaa
proteintranslation
AP Biology
____________ RNA structure “Clover leaf” structure
anticodon on “clover leaf” end amino acid attached on 3 end
AP Biology
Loading tRNA Aminoacyl tRNA synthetase
enzyme which bonds amino acid to tRNA bond requires energy
ATP AMP bond is unstable so it can release amino acid at ribosome easily
activatingenzyme
anticodontRNATrp binds to UGG
condon of mRNA
Trp Trp Trp
mRNAA C CU GG
C=O
OHOH
H2OO
tRNATrp
tryptophan attached to tRNATrp
C=O
O
C=O
AP Biology
Ribosomes Facilitate coupling of
tRNA anticodon to mRNA codon organelle or enzyme?
Structure ribosomal RNA (rRNA) & proteins 2 subunits
large small E P A
Protein synthesis/quiz
AP Biology
Ribosomes
Met
5'
3'
UUA C
A G
APE
A site (aminoacyl-tRNA site) holds tRNA carrying next amino acid to
be added to chain P site (peptidyl-tRNA site)
holds tRNA carrying growing polypeptide chain
E site (exit site) empty tRNA
leaves ribosome from exit site
Protein synthesis 2
AP Biology
Building a polypeptide ________________
brings together mRNA, ribosome subunits, initiator tRNA
_________________ adding amino acids based on
codon sequence
_________________ end codon 123
Leu
Leu Leu Leu
tRNA
Met MetMet Met
PE AmRNA5' 5' 5' 5'
3' 3' 3'3'
U UA AAACC
CAU UG G
GUU
A AAAC
CC
AU UG GGU
UA
AAAC
CC
AU UG GGU U
A AACCA U UG G
G AC
ValSer
AlaTrp
releasefactor
AA A
CCU UGG 3'
How translation works
AP Biology
Protein targeting ______________________
address label
Destinations: secretion nucleus mitochondria chloroplasts cell membrane cytoplasm etc…start of a secretory pathway
AP Biology
Can you tell the story?
DNA
pre-mRNA
ribosome
tRNA
aminoacids
polypeptide
mature mRNA
5' GTP cap
poly-A tail
large ribosomal subunit
small ribosomal subunit
aminoacyl tRNAsynthetase
E P A
5'
3'
RNA polymerase
exon intron
tRNA
AP Biology AAAAAAAAGTP
20-30b
3'
promoter transcriptionstop
transcriptionstart
introns
The Transcriptional unit (gene?)
transcriptional unit (gene)TAC ACT
DNA
DNATATA5'RNA
polymerase
pre-mRNA
5' 3'
translationstart
translationstop
mature mRNA
5' 3'
UTR UTR
exonsenhancer
1000+b
AP Biology 2007-2008
Protein Synthesis in Prokaryotes
Bacterial chromosome
mRNA
Cell wall
Cellmembrane
Transcription
Psssst…no nucleus!
AP 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
intronscome out!
AP Biology
Transcription & translation are simultaneous in bacteria DNA is in
cytoplasm no mRNA
editing ribosomes
read mRNA as it is being transcribed
Translation in Prokaryotes
AP Biology
Translation: prokaryotes vs. eukaryotes Differences between prokaryotes &
eukaryotes time & physical separation between
processes takes eukaryote ~1 hour
from DNA to protein no RNA processing
SEE PROCESSING VIDEO
COMPLETING PROTEINS POLYRIBOSOMES (POLYSOMES)
Numerous ribosomes translate same mRNA at same time
3-D folding (1’, 2’, 3’ structure) Chaparonins
POST-TRANSLATIONAL MODIFICATIONS Some amino acids modified by addition of
sugars, lipids, phosphate groups, etc Enzymes can modify ends, cleave into pieces
join polypeptide strands (4’ structure)
Ex: Made as proinsulin then cut
Final insulin hormonemade of two chains connected by disulfide bridges
http://www.vivo.colostate.edu/hbooks/pathphys/endocrine/pancreas/insulin.html
1st to suggest genes dictate phenotypes through enzymes that catalyze specific chemical reactions
Postulated that the symptoms of an inherited disease are due to inability to make a specific enzyme
Coined term “inborn errors of metabolism” to describe such diseases Beginning of “One gene-one enzyme” hypothesis ALCAPTONURIA- “black urine” disease- defect in enzyme that breaks down amino acid
tyrosine
http://www.personal.psu.edu/faculty/w/x/wxm15/Online/Molecular%20Biology/media/phenylalanine.gifhttp://www.nature.com/bjp/journal/v147/n1s/images/0706466f5.jpg
ARCHIBALD GARROD1902
Mutations 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
Slide from Explore Biology by Kim Foglia
Point mutation leads to Sickle cell anemia
What kind of mutation?
Slide from Explore Biology by Kim Foglia
Mutations
Frameshift shift in the reading
frame changes everything
“downstream” insertions
adding base(s) deletions
losing base(s) More damaging at
beginning of gene than at end
Slide modified from: Explore Biology by Kim Foglia
WHAT IS A “GENE”?
Mendel’s factors determine phenotype
T.H. Morgan- genes located on specific chromosomes
Beadle and Tatum’s “one gene-one enzyme”
Became “One gene-one polypeptide”
- Some proteins made of more than one polypeptide chainEx: hemoglobin has 4 polypeptide chains
Now: “one gene – one polypeptide or RNA” - Not all genes code for proteins
Recommended