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From Gene
to Protein
How Genes
Work
AP Biology
2007-2008
Making proteins

Organelles
nucleus
 ribosomes
 endoplasmic reticulum
(ER)
 Golgi apparatus
 vesicles

nuclear pore
small
ribosomal
subunit
mRNA
large
ribosomal
subunit
AP Biology
cytoplasm
Nucleolus

Function

ribosome production


build ribosome subunits from rRNA & proteins
exit through nuclear pores to cytoplasm &
combine to form functional ribosomes
large subunit
small
subunit
AP Biology
rRNA &
proteins
ribosome
nucleolus
large
subunit
Ribosomes

Function


small
subunit
protein production
Structure


rRNA & protein
2 subunits combine
0.08mm
Ribosomes
Rough
ER
Smooth
ER
AP Biology
Types of Ribosomes

Free ribosomes



suspended in cytosol
synthesize proteins that
function in cytosol
Bound ribosomes


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attached to endoplasmic
reticulum
synthesize proteins
for export or
for membranes
membrane proteins
TO:
endoplasmic
reticulum
nucleus
protein
on its way!
DNA
RNA
TO:
vesicle
TO:
TO:
vesicle
TO:
ribosomes
finished
protein
protein
Golgi
apparatus
Making Proteins
AP Biology
Metabolism taught us about genes

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
metabolic pathway
A

AP Biology enzyme 1
Am I just the
sum of my proteins?
disease
disease
disease
disease
B
C
D
E

enzyme 2

enzyme 3

enzyme 4
1941 | 1958
Beadle & Tatum
one gene : one enzyme hypothesis
George Beadle
Edward Tatum
AP Biology
"for their discovery that genes act by
regulating definite chemical events"
The “Central Dogma”

Flow of genetic information in a cell

How do we move information from DNA to proteins?
DNA
replication
AP Biology
RNA
protein
DNA gets
all the glory,
but proteins do
all the work!
trait
RNA


ribose sugar
N-bases
uracil instead of thymine
U : A
C : G



single stranded
lots of RNAs

DNA
AP Biology
mRNA, tRNA, rRNA, siRNA…
transcription
RNA
Transcription
from
DNA nucleic acid language
to
RNA nucleic acid language
AP Biology
2007-2008
Transcription

Making mRNA


transcribed DNA strand = template strand
untranscribed DNA strand = coding strand


synthesis of complementary RNA strand


same sequence as RNA
transcription bubble
enzyme

RNA polymerase
5
C
DNA
G
3
A
G
T
A T C
T A
53
G
A G C
A
T
C G T
A
C
T
3
G C A U C G U
C
G T A G C A
T
T
A
C
A G
C T
G
A
T
A
T
3
5
unwinding
rewinding
mRNA
AP Biology
build
RNA
coding strand
5
RNA polymerase
template strand
Transcription in Eukaryotes
Transcription
Psssst…
DNA can’t
leave nucleus!
RNA Processing
Translation
Protein
AP Biology
2007-2008
Prokaryote vs. Eukaryote genes

Prokaryotes

Eukaryotes


DNA in cytoplasm
circular
chromosome
naked DNA

no introns





DNA in nucleus
linear
chromosomes
DNA wound on
histone proteins
introns vs. exons
introns
come out!
intron = noncoding (inbetween) sequence
eukaryotic
DNA
exon = coding (expressed) sequence
AP Biology
Transcription in Eukaryotes

3 RNA polymerase enzymes

RNA polymerase 1



RNA polymerase 2


AP Biology
transcribes genes into mRNA
RNA polymerase 3


only transcribes rRNA genes
makes ribosomes
only transcribes tRNA genes
each has a specific promoter sequence
it recognizes
Transcription in Eukaryotes

Initiation complex

transcription factors bind
to promoter region
upstream of gene


suite of proteins which bind
to DNA
 turn on or off transcription
TATA box binding site
 recognition site for
transcription factors

transcription factors
trigger the binding of RNA
polymerase to DNA
AP Biology
Post-transcriptional processing

Primary transcript (pre-mRNA)


eukaryotic mRNA needs work after transcription
mRNA processing (making mature mRNA)


mRNA splicing = edit out introns
protect mRNA from enzymes in cytoplasm
 add 5 cap
mRNA
P
 add polyA tail
5' G PP
3'
A
intron = noncoding (inbetween) sequence
~10,000 bases
eukaryotic DNA
exon = coding (expressed) sequence
pre-mRNA
primary mRNA
transcript
AP Biology
mature mRNA
transcript
~1,000 bases
spliced mRNA
Splicing must be accurate

No room for mistakes!
splicing must be exactly accurate
 a single base added or lost throws off the
reading frame
AUGCGGCTATGGGUCCGAUAAGGGCCAU
AUGCGGUCCGAUAAGGGCCAU
AUG|CGG|UCC|GAU|AAG|GGC|CAU
Met|Arg|Ser|Asp|Lys|Gly|His

AP Biology
AUGCGGCTATGGGUCCGAUAAGGGCCAU
AUGCGGGUCCGAUAAGGGCCAU
AUG|CGG|GUC|CGA|UAA|GGG|CCA|U
Met|Arg|Val|Arg|STOP|
Whoa! I think
we just broke
a biological “rule”!
Splicing enzymes

snRNPs



small nuclear RNA
exon
proteins
Spliceosome


intron
3'
spliceosome
5'
3'
cut & paste
lariat
5'
AP Biology
exon
5'
several snRNPs
recognize splice
site sequence

snRNPs
snRNA
mature mRNA
exon
5'
3'
exon
3'
excised
intron
Ribozyme

1982 | 1989
RNA as ribozyme
some mRNA can even splice itself
 RNA as enzyme

Sidney Altman
AP Biology
Yale
Thomas Cech
U of Colorado
Translation
from
nucleic acid language
to
amino acid language
AP Biology
2007-2008
Translation

Codons

AP Biology
blocks of 3
nucleotides
decoded into
the sequence
of amino acids
Translation: prokaryotes vs. eukaryotes

Differences between prokaryotes &
eukaryotes

time & physical separation between
processes


AP Biology
takes eukaryote ~1 hour
from DNA to protein
RNA processing
Translation in Eukaryotes
AP Biology
2007-2008
From gene to protein
transcription
DNA
mRNA
mRNA leaves
nucleus through
nuclear pores
AP Biology
translation
nucleus
a
a
a
a
a
a
a
a
a
a
protein
a
ribosomea
a
a
a
a
proteins synthesized
by ribosomes using
instructions on mRNA
cytoplasm
How does mRNA code for proteins?
DNA
TACGCACATTTACGTACGCGG
4 ATCG
mRNA
4 AUCG
protein
AUGCGUGUAAAUGCAUGCGCC
?
Met Arg Val Asn Ala Cys Ala
20
AP Biology
How can you code for 20 amino acids
with only 4 nucleotide bases (A,U,G,C)?
mRNA codes for proteins in triplets
DNA
TACGCACATTTACGTACGCGG
codon
mRNA
AUGCGUGUAAAUGCAUGCGCC
?
protein
AP Biology
Met Arg Val Asn Ala Cys Ala
Cracking the code

1960 | 1968
Nirenberg & Khorana
Crick

determined 3-letter (triplet) codon system
WHYDIDTHEREDBATEATTHEFATRAT

Nirenberg (47) & Khorana (17)
determined mRNA–amino acid match
 added fabricated mRNA to test tube of
ribosomes, tRNA & amino acids



AP Biology
created artificial UUUUU… mRNA
found that UUU coded for phenylalanine (phe)
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”
Why is the
wobble good?

Start codon



AP Biology
AUG
methionine
Stop codons

UGA, UAA, UAG
How are the codons matched to
amino acids?
DNA
mRNA
3
5
5
3
TACGCACATTTACGTACGCGG
AUGCGUGUAAAUGCAUGCGCC
codon
3
tRNA
UAC
amino
acid
Met
AP Biology
5
GCA
Arg
CAU
Val
anti-codon
From gene to protein
transcription
DNA
translation
mRNA
a
ribosomea
a
a
a
a
a
a
a
a
a
a
protein
a
a
a
a
aa
AP Biology
nucleus
cytoplasm
Transfer 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
energy stored in tRNA-amino acid bond


unstable
so it can release amino acid at ribosome easily
Trp C=O
OH
OH
Trp C=O
O
Trp
H2O
O
activating
enzyme
tRNATrp
anticodon
AP Biology
tryptophan attached
to tRNATrp
AC C
UGG
mRNA
tRNATrp binds to UGG
condon of mRNA
Ribosomes

Facilitate coupling of
tRNA anticodon to
mRNA codon


organelle or enzyme?
Structure
ribosomal RNA (rRNA) & proteins
 2 subunits



AP Biology
large
small
E P A
Ribosomes

A site (aminoacyl-tRNA site)


P site (peptidyl-tRNA site)


holds tRNA carrying next amino acid to
be added to chain
holds tRNA carrying growing
polypeptide chain
Met
E site (exit site)

AP Biology
empty tRNA
leaves ribosome
from exit site
U A C
A U G
5'
E
P
A
3'
Building a polypeptide

Initiation


Elongation


brings together mRNA, ribosome
subunits, initiator tRNA
adding amino acids based on
codon sequence
Termination

3 2 1
end codon
Leu
Val
Met
Met
Met
Met Leu
Ala
Leu
Leu
release
factor
Ser
Trp
tRNA
U AC
5'
C UGAA U
mRNA A U G
3'
E P A
AP Biology
5'
UAC GAC
A U G C U GAA U
5'
3'
U A C GA C
A U G C U G AAU
5'
3'
U AC G A C
AA U
AU G C U G
3'
A CC
U GG U A A
3'
Destinations:
Protein targeting



Signal peptide

address label




start of a secretory pathway
AP Biology

secretion
nucleus
mitochondria
chloroplasts
cell membrane
cytoplasm
etc…
RNA polymerase
DNA
Can you tell
the story?
amino
acids
exon
intron
tRNA
pre-mRNA
5' cap
mature mRNA
aminoacyl tRNA
synthetase
polyA tail
large ribosomal subunit
polypeptide
5'
small ribosomal subunit
AP Biology
tRNA
E P A
ribosome
3'
Got Questions?
Can I translate that for you?
AP Biology
2007-2008
Bacterial chromosome
Transcription
in Prokaryotes
Transcription
mRNA
Psssst…
no nucleus!
Cell
membrane
Cell wall
AP Biology
2007-2008
Transcription in Prokaryotes

Initiation

RNA polymerase binds to promoter
sequence on DNA
Role of promoter


Starting point
 where to start reading
 start of gene
Template strand
 which strand to read
Direction on DNA
 always read DNA 35
APBiology
build RNA 53

Transcription in Prokaryotes

Promoter sequences
enzyme
subunit
RNA polymerase
read DNA 35
bacterial DNA
Promoter
TTGACA TATAAT
–35 sequence
–10 sequence
RNA polymerase
molecules bound to
bacterial DNA
RNA polymerase
AP Biology
strong vs. weak promoters
Transcription in Prokaryotes

Elongation

RNA polymerase
copies DNA as it
unwinds


~20 base pairs at a time
 300-500 bases in gene
builds RNA 53
Simple proofreading
1 error/105 bases
 make many mRNAs
 mRNA has short life
not worth editing!
AP 
Biology

reads DNA 35
Transcription in Prokaryotes

Termination

RNA polymerase stops at termination
sequence
RNA GC
hairpin turn
AP Biology
Bacterial chromosome
Translation in
Prokaryotes
Transcription
mRNA
Translation
Psssst…
no nucleus!
protein
Cell
membrane
Cell wall
AP Biology
2007-2008
Translation in Prokaryotes

Transcription & translation are simultaneous
in bacteria
DNA is in
cytoplasm
 no mRNA
editing
 ribosomes
read mRNA
as it is being
transcribed

AP Biology