Download Question How does DNA control a cell?By controlling Protein

Chapter 17:
From Gene to Protein
(Protein Synthesis)
Essential Knowledge
3.a.1 – DNA, and in some cases RNA, is
the primary source of heritable
information (17.1-17.4).
 3.c.1 – Changes in genotype can result
in changes in phenotype (17.5).

Question?

How does DNA control a cell? (or identify a
phenotype)

By controlling protein synthesis (otherwise
known as gene expression)

Proteins are the link between genotype and
phenotype
1909 - Archibald Garrod
Suggested genes control enzymes that
catalyze chemical processes in cells
 Inherited Diseases - “inborn errors of
metabolism” where a person can’t make an
enzyme

 Symptoms reflect person’s inability to make proteins/enzymes
Example
Alkaptonuria - where urine turns black
after exposure to air
 Lacks - an enzyme to metabolize/break
down alkapton

George Beadle and Edward
Tatum
Worked with Neurospora and proved the
link between genes and enzymes
 Grew Neurospora on agar
 Varied the nutrients in the agar
 Looked for mutants that failed to grow
on minimum agar

Conclusion
Mutations were abnormal genes
 Each gene dictated the synthesis/production
of one enzyme
 One Gene - One Enzyme Hypothesis

Current Hypothesis

One Gene - One Polypeptide
Hypothesis.
 Why change? Not all proteins are enzymes

We now know proteins may have 4th
degree structure.
Central Dogma
DNA
Transcription
RNA
Translation
Polypeptide chain
(will become protein)
Explanation
DNA – the genetic code or genotype
 RNA - the message or instructions
 Polypeptide - the end product for the
phenotype

Why is there an RNA
intermediate?

Evolutionary adaptation:
 Check-point in process
 Provides protection for DNA code
 More copies can be made simultaneously
Genetic Code
Sequence of DNA bases that describe which
amino acid to place in what order in a
polypeptide chain
 The genetic code gives ONLY the primary
protein structure

 All other protein structures result from chemical interactions
amongst primary protein structure
Genetic Code
 Is
based on triplets of bases (called
codons)
 Has redundancy; some AA's have more
than 1 code/3-base codon
 Proof - make artificial RNA and see what
AAs are used in protein synthesis (early
1960’s)
Codon
A 3-nucleotide “word” in the Genetic
Code
 64 possible codons known

Codon
Amino
acid
Codon Dictionary
Start- AUG (Met)
 Stop- UAA
UAG
UGA
 60 codons for the other 19 AAs

Code Redundancy
 Third
base in a codon shows "wobble”
effect
 First two bases are the most important in
reading the code and giving the correct
AA
 The third base often doesn’t matter
 This allows for mistakes during DNA
replication
Reading Frame
 The
“reading” of the code is every three
bases
 Ex: the red cat ate the rat
 Ex: ATT GAT TAC ATT
 The
“words” (codons) only make sense
if “read” in this grouping of three (in
correct “letter” order)
Code Evolution
The genetic code is nearly universal
 Ex: CCG = proline (all life)
 Reason:

 Code must have evolved early
 Life on earth must share a common ancestor
Protein Synthesis Intro

Intro movie
Protein Synthesis Intro

Step 1: Transcription:
 DNA  mRNA

Step 2: Translation:
 mRNA  tRNA  Am. Acid  Polypep.
chain

Polypeptide chain then becomes protein
Transcription

Process of making RNA from a DNA
template
 RNA type: mRNA (messenger)
 Intermediate type

Takes place in nucleus (in eukaryotes)
Transcription Steps
1.
2.
3.
4.
RNA Polymerase Binding
Initiation
Elongation
Termination
RNA Polymerase
 Enzyme
for building RNA from RNA
nucleotides
 Prokaryotes - 1 type
 Eukaroyotes- 3 types
 Splits
two DNA strands apart
 Hooks RNA nucleotides together (as they
pair with DNA)
st
1
Step: RNA Polymerase
Binding
Requires that the enzyme find the
“proper” place on the DNA to attach and
start transcription
 Different from DNA polymerase

 Doesn’t require an RNA primer
RNA Polymerase Binding Needs:
 Promoter
Regions (on the DNA)
 Special sequences of DNA nucleotides that
“tell” cell where transcription begins
 Transcription
 Proteins
Factors
Promoters
Regions of DNA where RNA Polymerases can
bind
 About 100 nucleotides long. Include initiation
site and recognition areas for RNA Polymerase
 Also “decide” which DNA strand to use

TATA Box
ONLY in eukaryotes
 Short segment of T,A,T,A
 Located 25 nucleotides upstream from the
initiation site
 Recognition site for transcription factors to bind
to the DNA

Transcription Factors
Proteins that bind to DNA before RNA
Polymerase
 Recognizes TATA box, attaches, and
“flags” the spot for RNA Polymerase

 RNA poly won’t attach unless these are
present
Transcription Initiation Complex

The complete assembly of
 1) transcription factors and
 2) RNA Polymerase

Bound to the promoter area of the DNA to
be transcribed
nd
2
Step: Initiation
2nd step of transcription
 Actual unwinding of DNA to start RNA
synthesis.
 Requires Initiation Factors

Comment
Getting Transcription started is
complicated
 Gives many ways to control which
genes are decoded and which proteins
are synthesized

rd
3
Step: Elongation
3rd step in transcription
 RNA Polymerase untwists DNA 1 turn at a time
 Exposes 10 DNA bases for pairing with RNA
nucleotides

Elongation
Adds nucleotides to 3` end of growing
RNA strand
 Enzyme moves 5`  3` (of RNA strand)
 Rate is about 60 nucleotides per second

Comment
Each gene can be read by sequential
RNA Polymerases giving several copies
of RNA
 Result - several copies of the protein
can be made

th
4
Step: Termination
DNA sequence that tells RNA Polymerase
to stop
 Ex: AATAAA
 RNA Polymerase detaches from DNA
after closing the helix

Final Product
Pre-mRNA
 This is a “raw” RNA that will need
processing and modifications

Modifications of RNA
1. 5’ Cap
2. Poly-A Tail
3. Splicing
5' Cap
Modified Guanine nucleotide added to
the 5' end
 Protects mRNA from digestive enzymes
 Recognition sign for ribosome
attachment

Poly-A Tail
150-200 Adenine nucleotides added to
the 3' tail
 Protects mRNA from digestive enzymes.
 Aids in mRNA transport from nucleus.

RNA Splicing
Removal of non-protein coding regions
of RNA
 Coding regions are then spliced back
together

Introns and Exons

Introns:
 Intervening sequences
 Removed from RNA.

Exons:
 Expressed sequences of RNA
 Translated into AAs
Introns - Function
Left-over DNA (?)
 Way to lengthen genetic message
 Old virus inserts (?)
 Way to create new proteins

Translation Poster
Requirements
1. What is translation? (definition)
 2. What is needed?
 3. Specifics & Structure of tRNA
 4. Where does it occur?
 5. Ribosome specifics – be sure to include the
specifics of each subunit
 6. Steps of translation & details of each step
 7. What bonds are formed?
 8. Illustration

2nd step of Protein
Synthesis: Translation
Process by which a cell interprets a
genetic message and builds a
polypeptide
 Location: mRNA moves from nucleus to
cytoplasm and ribosomes

Materials Required for
translation
tRNA
 Ribosomes
 mRNA

Transfer RNA = tRNA
Made by transcription
 About 80 nucleotides long
 Carries AA for polypeptide synthesis

Structure of tRNA
Has double stranded regions and 3
loops.
 AA attachment site at the 3' end.
 1 loop serves as the Anticodon.

Anticodon
Region of tRNA that base pairs to mRNA codon
 Usually is a compliment to the mRNA bases, so
reads the same as the DNA codon
 Example:

 DNA- GAC
 mRNA – CUG
 tRNA anticodon - GAC
Ribosomes
Two subunits (large and small) made in
the nucleolus
 Made of rRNA (60%)and protein (40%)
 rRNA is the most abundant type of RNA
in a cell

Large Subunit
Has 3 sites for tRNA.
 P site: Peptidyl-tRNA site - carries the
growing polypeptide chain
 A site: Aminoacyl-tRNA site -holds the tRNA
carrying the next AA to be added
 E site: Exit site

Translation Steps
1. Initiation
2. Elongation
3. Termination
Initiation

Brings together:
 mRNA
 A tRNA carrying the 1st AA
 2 subunits of the ribosome
Initiation Steps:
1. Small subunit binds to the
mRNA
2. Initiator tRNA (Met, AUG)
binds to mRNA
3. Large subunit binds to mRNA
Initiator tRNA is in the P-site
Initiation
Requires other proteins called "Initiation
Factors”
 GTP used as energy source

Elongation Steps:
1. Codon Recognition
2. Peptide Bond Formation
3. Translocation
Codon Recognition

tRNA anticodon matched to mRNA
codon in the A site
Peptide Bond Formation
A peptide bond is formed between the new
AA and the polypeptide chain in the P-site
 Bond formation is by rRNA acting as a
ribozyme
 After bond formation:

 The polypeptide is now transferred from the tRNA
in the P-site to the tRNA in the A-site
Translocation
tRNA in P-site is released
 Ribosome advances 1 codon, 5’ 3’
 tRNA in A-site is now in the P-site
 Process repeats with the next codon

Termination
Triggered by stop codons
 Release factor binds in the A-site
instead of a tRNA
 H2O is added instead of AA, freeing the
polypeptide
 Ribosome separates

Polyribosomes
Cluster of ribosomes all reading the
same mRNA
 Another way to make multiple copies of
a protein

Prokaryotes: Prok. vs. Euk. Protein
Synthesis Video
Polypeptide vs. Protein
Polypeptide usually needs to be
modified before it becomes functional
 Ex:

 Sugars, lipids, phosphate groups added
 Some AAs removed
 Protein may be cleaved
 Join polypeptides together (Quaternary
Structure)
Mutations
Changes in the genetic make-up of a
cell
 Chapter 15 covered large-scale
chromosomal mutations

 (Hint - review these!)
Mutation types - Cells
Somatic cells or body cells – not
inherited
 Germ Cells or gametes - inherited

Point or Spot Mutations
Changes in one or a few nucleotides in
the genetic code
 Effects - none to fatal

Types of Point Mutations
1. Base-Pair Substitutions
2. Insertions
3. Deletions
Base-Pair Substitution
The replacement of 1 pair of nucleotides
by another pair
 Ex: Sickle cell anemia

Types of Substitutions
1. Missense - altered codons, still code for
AAs but not the right ones
2. Nonsense - changed codon becomes a
stop codon
Question?
What will the "Wobble" Effect have on
Missense?
 If the 3rd base is changed, the AA may
still be the same and the mutation is
“silent”

Missense Effect
Can be none to fatal depending on
where the AA was in the protein
 Ex:

 If in an active site - major effect
 If in another part of the enzyme - no effect
Nonsense Effect
Stops protein synthesis
 Leads to nonfunctional proteins unless
the mutation was near the very end of
the polypeptide

Sense Mutations
The changing of a stop codon to a
reading codon
 Result - longer polypeptides which may
not be functional
 Ex. “heavy” hemoglobin

Insertions & Deletions
The addition or loss of a base in the
DNA
 Cause frame shifts and extensive
missense, nonsense or sense mutations

Frame Shift
The “reading” of the code is every three
bases
 Ex: the red cat ate the rat
 Ex: thr edc ata tat her at
 The “words” only make sense if “read” in this
grouping of three

Question?
Loss of 3 nucleotides is often not a
problem
 Why?

 Because the loss of a 3 bases or one codon
restores the reading frame
Mutagens
Materials that cause DNA changes
1. Radiation

ex: UV light, X-rays
2. Chemicals
ex: 5-bromouracil
Summary
Recognize the relationship between genes
and enzymes (proteins) as demonstrated by
the experiments of Beadle and Tatum.
 Identify the flow of genetic information from
DNA to RNA to polypeptide (the “Central
Dogma”).
 Read DNA or RNA messages using the
genetic code.
 Recognize the steps and procedures in
transcription.

Summary
 Identify
methods of RNA modification.
 Recognize the steps and procedures
in translation.
 Recognize categories and
consequences of base-pair mutations.
 Identify causes of mutations.
 Be able to recognize and discuss
“What is a gene?”