Download MCDB 1041 3/9/12 Activity 6: Central Dogma Continued PART I

MCDB 1041 3/9/12
Activity 6: Central Dogma Continued
PART I: Types of mutations:
• Point mutations: changes in a single base of DNA.
Silent mutations: change in sequence of DNA results in no change in the amino acid sequence.
Missense mutations: change in DNA sequence results in a different amino acid placed into the protein.
Nonsense mutations: change in DNA creates a premature stop codon.
• Deletions or Insertions: removal or addition of 1 or more bases. The possible consequences depend
on the number of bases removed, and the position of their removal.
Frame-shift mutations: change in DNA shifts the reading frame, resulting in a completely
different amino acid sequence in the protein after the mutation. This can be the result of a deletion or an
insertion of 1 or 2 bases, or any multiple of 2 bases. Insertions or deletions of 3 or multiples of 3 do not
result in a frame shift. Why?
• Expanding repeat mutations: an error in copying the DNA results in an already repeated sequence
becoming longer (more repeats). This is like the STRs. Expanding repeats that happen within genes
do have an effect on phenotype (unlike STRs).
A sequence of amino acids coded for by an mRNA= a protein. Proteins have specific structures
dictated by bonds that form between the amino acids, giving the protein a specific shape, and allowing
it to perform specific functions. Thus, mutations in the DNA affect the structure of the protein (longer,
shorter, can’t fold properly, different sequence of amino acids, etc).
Reviewing Translation: the effects of different mutations on a protein
Ever noticed this label on cans of diet soda?
What does it mean?
What is phenylalanine?
Why can’t you eat aspartame when you have Phenylketonuria?
Phenylketonuria (PKU) is an autosomal recessive disorder caused by mutations in the DNA that codes for
an enzyme called phenylalanine hydroxylase. This enzyme breaks down the amino acid phenylalanine, which
is important because phenylalanine is toxic when it accumulates in nerve cells, and can cause brain damage in
children. Most proteins contain some of the amino acid phenylalanine, so people with PKU cannot each much
protein, and in particular, should never take in phenylalanine itself. The artificial sweetener Aspartame is
composed of two amino acids: phenylalanine and aspartic acid.
PKU can be caused by a number of different mutations in the phenylalanine hydroxylase gene. Different
mutations affect the activity of the enzyme to different extents, leading to more or less severe forms of PKU.
Below there are some actual alleles of the gene for phenylalanine hydroxylase enzyme. These are all
different regions of the gene, starting with the beginning of the gene in part A and continuing along the
extent of the gene. Thus, you will notice that only the sequence in part A begins with ATG, the required
start codon. The promoter would be just to the left of this ATG sequence (ie, the top strand is the coding
strand). Your tasks:
* Find and mark where the nucleotide change has occurred between the normal and mutant DNA sequence.
* Use the codon table to determine how the mutation in the DNA will affect the amino acid sequence. In
some cases, you may need to look at more than just the single change…because it may affect other codons!
A. Allele A, mutation found in French and French Canadian population
normal
5-ATG TCC ACT GCG GTC CTG GAA AAC CCA GGC-3
3-TAC AGG TGA CGC CAG GAC CTT TTG GGT CCG-5
Allele A
.
5-GTG TCC ACT GCG GTC CTG GAA AAC CCA GGC-3
3-CAC AGG TGA CGC CAG GAC CTT TTG GGT CCG-5
B. Allele B, Multiple ethnic associations.
Write out the entire amino acid sequence for the normal
and mutant alleles
5-TTA TTT GAG GTA AAG AAT GAT GAC-3
normal
3-AAT AAA CTC CAT TTC TTA CTA CTG -5
Allele B
5-TTA TTG AGG TAA AGA ATG ATG ACA -3
3-AAT AAC TCC AAT TCT TAC TAC TGT -5
C. Allele C, Identified in Swiss patients.
normal
5-GGT GGC CTG GCC TTC CGA GTC TTT-3
3-CCA CCG GAC CGG AAG GCT CAG AAA-5
Allele C
5-GGT GGC CTG GCC TTC CAA GTC TTT-3
3-CCA CCG GAC CGG AAG GTT CAG AAA-5
Kind of mutation:
What affect does the
mutation have on the
transcription?
What affect is this
mutation likely to have on
the protein?
Kind of mutation:
What affect does the
mutation have on the
transcription?
What affect is this
mutation likely to have on
the protein?
Kind of mutation:
What affect does the
mutation have on the
transcription?
What affect is this
mutation likely to have on
the protein?
D. Allele D, Identified in Chinese patients.
normal
Allele D
5-AAG GAG AAA GTA AGG AAC TTT GCT GCC ACA-3
3-TTC CTC TTT CAT TCC TTG AAA CGA CGG TGT-5
5-AAG GAG AAA GTT AGG AAC TTT GCT GCC ACA-3
3- TTC CTC TTT CAA TCC TTG AAA CGA CGG TGT-5
Kind of mutation:
What affect does the
mutation have on the
transcription?
What affect is this
mutation likely to have
on the protein?
Of these four mutations, which one do you think will cause the most
severe symptoms of PKU (ie, in which case will the protein be most dysfunctional?)
PART II. Putting it all together
1. As a group use a series of analogies to capture the essence of the central dogma. You should include
analogies for DNA replication, RNA transcription, and Translation, as well as including RNA polymerase,
the ribosome, and the tRNA.
2. Now, create a concept map of the different terms you have used to describe gene expression. A concept
map works like this: using the terms below, arrange them into a map, where a phrase connects each term
with a line. For example, the term “DNA” could be connected with a line and the words “is composed of”
to another term “nucleotides”. Do the concept map together as a group, on the white boards, and then copy
it down for yourself here.
Terms:
DNA
Replication
DNA polymerase
Template strand
Coding strand
mRNA
RNA polymerase
Promoter region
Unique transcription factors
Ribosome
tRNA
amino acids
alternative splicing
cytoplasm
nucleus