chapter 15 from genes to proteins. question? u how does dna control a cell? u by controlling protein...
TRANSCRIPT
Question?
How does DNA control a cell? By controlling Protein
Synthesis. Proteins are the link between
genotype and phenotype.
For tests:
Name(s) of experimenters Outline of the experiment Result of the experiment and
its importance
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.
Example
Alkaptonuria - where urine turns black after exposure to air.
Lacks - an enzyme to metabolize alkapton.
George Beadle and Edward Tatum
Worked with Neurospora and proved the link between genes and enzymes.
Neurospora
Pink bread mold
Experiment
Grew Neurospora on agar. Varied the nutrients. Looked for mutants that
failed to grow on minimum agar.
Results
Three classes of mutants for Arginine Synthesis.
Each mutant had a different block in the Arginine Synthesis pathway.
Conclusion
Mutations were abnormal genes.
Each gene dictated the synthesis of one enzyme.
One Gene - One Enzyme Hypothesis.
Explanation
DNA - the Genetic code or genotype.
RNA - the message or instructions.
Polypeptide - the product for the phenotype.
Genetic Code
Sequence of DNA bases that describe which Amino Acid to place in what order in a polypeptide.
The genetic code gives the primary protein structure.
Code Basis
If you use: 1 base = 1 amino acid 4 bases = 4 amino acids 41 = 4 combinations, which
are not enough for 20 AAs.
Genetic Code
Is based on triplets of bases. Has redundancy; some AA's
have more than 1 code. Proof - make artificial RNA and
see what AAs are used in protein synthesis (early 1960’s).
For Testing:
Be able to “read” a DNA or RNA message and give the AA sequence.
RNA Genetic Code Table will be provided.
Code Redundancy
Third base in a codon shows "wobble”.
First two bases are the most important in reading the code and giving the correct AA. The third base often doesn’t matter.
Code Evolution
The genetic code is nearly universal.
Ex: CCG = proline (all life) Reason - The code must have
evolved very early. Life on earth must share a common ancestor.
Reading Frame and Frame Shift
The “reading” of the code is every three bases (Reading Frame)
Ex: the red cat ate the rat Frame shift – improper groupings
of the bases Ex: thr edc ata tat her at The “words” only make sense if
“read” in this grouping of three.
Binding
Requires that the enzyme find the “proper” place on the DNA to attach and start transcription.
Binding
Is a complicated process Uses Promoter Regions on
the DNA (upstream from the information for the protein)
Requires proteins called Transcription Factors.
Transcription Initiation Complex
The complete assembly of transcription factors and RNA Polymerase bound to the promoter area of the DNA to be transcribed.
Elongation
RNA Polymerase untwists DNA 1 turn at a time.
Exposes 10 DNA bases for pairing with RNA nucleotides.
Comment
Each gene can be read by sequential RNA Polymerases giving several copies of RNA.
Result - several copies of the protein can be made.
Termination
DNA sequence that tells RNA Polymerase to stop.
Ex: AATAAA RNA Polymerase detaches
from DNA after closing the helix.
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.
Let’s see Transcription in motion…
http://www.hhmi.org/biointeractive/media/DNAi_transcription_vo2-lg.mov
RNA Splicing
Removal of non-protein coding regions of RNA.
Coding regions are then spliced back together.
Ribozymes
RNA molecules that act as enzymes.
Are sometimes Intron RNA and cause splicing without a spliceosome.
Introns - Function
Left-over DNA (?) Way to lengthen genetic
message. Old virus inserts (?) Way to create new proteins.
Alternative Splicing
The RNA can be spliced into different mRNA’s.
Each different mRNA produces a different polypeptide.
Ex. – variable regions of antibodies.
DSCAM Gene
Found in fruit flies Has 100 potential splicing sites. Could produce 38,000 different
polypeptides Many of these polypeptides have
been found
Commentary
Alternative Splicing is going to be a BIG topic in Biology.
About 60% of genes are estimated to have alternative splicing sites.
One gene does not equal one polypeptide.
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.
Comment
"Wobble" effect allows for 45 types of tRNA instead of 61.
Reason - in the third position, U can pair with A or G.
Inosine (I), a modified base in the third position can pair with U, C, or A.
Importance
Allows for fewer types of tRNA.
Allows some mistakes to code for the same AA which gives exactly the same polypeptide.
Aminoacyl-tRNA Synthetases
Family of Enzymes. Add AAs to tRNAs. Active site fits 1AA and 1 type of
tRNA. Uses a “secondary genetic” code
to load the correct AA to each tRNA.
Ribosomes
Two subunits 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
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
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.
Let’s see Translation in motion…
http://www.hhmi.org/biointeractive/media/DNAi_translation_vo2-lg.mov
Polyribosomes
Cluster of ribosomes all reading the same mRNA.
Another way to make multiple copies of a protein.
Examples
Sugars, lipids, phosphate groups added.
Some AAs removed. Protein may be cleaved. Join polypeptides together
(Quaternary Structure).
Signal Hypothesis
“Clue” on the growing polypeptide that causes ribosome to attach to ER.
All ribosomes are “free” ribosomes unless clued by the polypeptide to attach to the ER.
Result
Protein is made directly into the ER .
Protein targeted to desired location (e.g. secreted protein).
“Clue” (the first 20 AAs are removed by processing).
General Result
Loss of genetic information. Position effects: a gene's
expression is influenced by its location to other genes.
Cri Du Chat Syndrome
Part of p arm of #5 has been deleted.
Good survival. Severe mental retardation. Small sized heads common.
Philadelphia Chromosome
An abnormal chromosome produced by a translocation of portions of chromosomes 9 and 22.
Causes chronic myeloid leukemia.
Mutation types - Cells
Somatic cells or body cells – not inherited
Germ Cells or gametes - inherited
DNA or Point Mutations
Changes in one or a few nucleotides in the genetic code.
Effects - none to fatal.
Lets see how this mutation will affect the cell…
http://www.hhmi.org/biointeractive/media/DNAi_sicklecell-lg.mov
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.
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 and the protein may still be able to function.
Mutagens
Materials that cause DNA changes.
1. Radiationex: UV light, X-rays
2. Chemicalsex: 5-bromouracil
Comment
Any material that can chemically bond to DNA, or is chemically similar to the nitrogen bases, will often be a very strong mutagen.
Summary Know Beadle and Tatum. Know the central dogma. Be able to “read” the genetic
code. Be able to describe the events
of transcription and translation.