MCB 150 Week 5 topic 1: mutations

What does proofreading and what direction does it go in?

3' ---> 5' exonuclease activity of DNA polymerase synthesis adds bases in 5' --> 3' direction last base added is checked if incorrect, last base is removed from 3' end

How often do errors occur in DNA replication?

Average in all organisms is 1 mistake in 1 billion nucleotides added

if proofreading doesn't catch the mistake...

Mismatch repair system of enzymes scans recently synthesized DNA: New DNA gets methylated at adenine residues within the sequence 5'-GATC-3' ~10 minutes after replication Brief period of time where parent strand is methylated and daughter strand is not: called Hemimethylated DNA

Which one of the following types of mutations would disrupt the reading frame in mRNA? Chemical-induced change of cytosine to thymine Same sense mutation Deletion of 1 or 2 bases Insertion of 3 bases Missense mutation

deletion of 1 or 2 bases

Duchenne's Muscular Dystrophy

deletion of exons in dystrophin: muscle fibers degeneration of muscle tissue

What happens if you strip away DNA polymerase's exonuclease activity?

it is 100 times more likely to end up introducing mutations than if that exonuclease activity, that proofreading activity, is there. And this accounts for half of that reduction in the observed overall error frequency. So, 100 times more likely to correct a mistake if you are doing proofreading.

Multi cellular organism germ-line mutation

sex cell: passed to new organism. affects survivability of the species: embryo will get mutation

base substitution mutation (2 types)

substitution of one base pair for another in DNA. because of the degenerate nature of DNA, this may or may not alter the amino acid encoded. 2 types: 1. silent mutation: new codon from base substitution still encodes the same amino acid 2. missense mutation: base substitution changes an amino acid

Cystic Fibrosis

Cystic Fibrosis is characterized by faulty protein (CFTR) 70 percent of CF cases are due to an in-frame deletion of a Phe codon at AA 508 of a 1,400 AA protein Mutant CFTR protein doesn't fold properly, never becomes an active channel Causes Cl- to accumulate in cells, causing water to be taken into cells This causes mucus glands to produce thick, sticky mucus, which interferes with breathing and digestion natural selection hasnt eliminated it because in certain population under certain circumstances it made selective advantage: carriers can resist cholera and survive because retaining more water in their cells

how does E. coli (or any other organism) reduce the frequency of the errors it makes by 104? (2 ways)

DNA polymerases can proofread their work (check as they go) Mismatch repair (spell-checker): happens right after Both occur during or quickly after DNA replication

Which of the processes that are dependent on correct base pairing is MOST reliable and why?

DNA replication because it is the blueprint for everything else and RNA are temporary

Match the genetic disorder to the type of mutation. [Note: each type of mutation may be used once, more than once, or not at all] A Insertion B Loss of an amino acid C Nonsense D Missense E Deletion Duchenne's muscular dystrophy Cystic fibrosis Fragile X syndrome Huntington's disease Sickle cell anemia

Duchenne's muscular dystrophy: E Cystic fibrosis: B Fragile X syndrome: A Huntington's disease: A Sickle cell anemia: D

Missense mutation affect on proteins...

IF... the AA that has been altered is not critical for the folding or function of that protein... OR... if the AA that replaces it assumes those duties in roughly the same way... (both acidic/negatively charged or basic/positively charged) THEN... the protein is likely to retain at least some function, may even be fully functional. BUT... if the altered AA is critical, and the duties aren't taken up by the AA that replaces it... THEN... the protein will be inactive.

Which one of the following statements about base substitution mutations is correct? Nonsense mutations are also called "silent" mutations. In same-sense mutations, usually the third nucleotide in a sequence is changed, resulting in a different codon for the same amino acid. If a base substitution mutation occurs in a protein-coding region of a gene, the resulting protein will always be non-functional. If a base substitution mutation occurs in a protein-coding region of a gene, the resulting protein will always be fully functional. Base substitution mutations occur when large pieces of DNA are inserted into a gene sequence.

In same-sense mutations, usually the third nucleotide in a sequence is changed, resulting in a different codon for the same amino acid.

How does repair mechanism know which strand to fix?

It's going to look for the strand in hemimethylated DNA that isn't yet methylated. Once DNA gets methylated, enzymes have no way of distinguishing parent from daughter strand: once window closes, it wouldn't even try to guess

Why are mistakes in Transcription or Translation not as critical?

Many copies of RNA are produced and RNAs are not heritable over multiple generations.

MutH, MutS, and MutL

MutH: looking for hemimethylated DNA and when it finds it it latches on and holds on for a second and waits to see if it gets a signal. once activated it causes a nick that recruits an exonuclease. That exonuclease is going to come in and strip out the region of DNA in the non-methylated strand all the way around the bend past the problem, just past the problem. MutS scans DNA looking for a problem (TG base pairs,etc). doesn't determine which one is wrong but it only finds theres a problem and forms a complex with MutL MutS + MutL: form a complex that links MutL/MutS with MutH and loops out whatever region of DNA lies between the problem and the nearest hemimethylated DNA sequence

In the mismatch repair system in E. coli, which one of the following enzymes does not directly perform a function? Primase MutH Exonucleases Ligase MutL

Primase

True or False... the negatively charged, positively charged, and polar amino acids are hydrophilic so they like being on the outside of an amino acid chain

TRUE

Determine if each of the following is a possible outcome of a protein coding gene with a missense mutation (and only that missense mutation). If the outcome is a possibility, choose Yes. If the outcome is not a possibility, choose No. The protein is prematurely truncated. The protein is fully functional. The protein is completely inactive. The protein has partial function.

The protein is prematurely truncated. NO The protein is fully functional. YES The protein is completely inactive. YES The protein has partial function. YES

Can one amino acid actually make a difference? give example using sickle cell anemia and how it works

Yes. Sickle cell anemia caused by defective beta globin subunit: changes overall charge of protein with distorted hemoglobin molecule so cells get stuck in blood vessels and limits oxygen delivery causing pain creates sticky sites causing hemoglobin molecules to stick together and precipitate out. pushes red blood cell outward with distorted shape 8% of African Americans are carriers. Why hasn't natural selection gotten rid of it? Parasite infects red blood cells which prevents malaria

point mutation

a mutation that alters a single base. can be either the substitution of one base for another or deletion or addition of a single base (or small number of bases)

slipped strand mispairing

a nucleotide sequence is present in more than one place on chromosome and copies in homologous chromosomes pair in wrong place. creates a loop in the chromosome

missense mutation (2 classes)

a type of mutation that changes an amino acid in a protein 2 classes: 1.transitions: doesn't change the type of base in the base pair (pyrimidine stays pyrimidine, purine stays purine) 2. transversion:does change the type of base pair.

frameshift mutation

addition or deletion of 1 or 2 bases. much worse consequences than a substitution of a base: affects everything downstream from it almost always results in premature stop codon

in single celled organisms, how many of the daughter cells have the mutation?

all of the daughter cells

Removing an in-frame, 3'-5' ATC sequence in the template strand of the coding region of a protein-coding gene would result in: a true frameshift mutation. an in-frame transition. an insertion mutation. an elongated polypeptide chain. a deletion mutation.

an elongated polypeptide chain

Why do most mistakes occur during DNA replication?

because were synthesizing huge amounts of DNA

Non sense mutation

change in base results in premature stop codon. protein truncated and virtually inactive (null phenotype)

DNA polymerase 1 has the ability to ....

do synthesis in the 5' to 3' direction primer removal by a 5' to 3' exonuclease activity proofreading by a 3' to 5' exonuclease activity This one enzyme has three different active sites in it. DNA polymerase 3 and most of the other DNA polymerases simply have the ability to do synthesis in the 5' to 3' direction and proofreading via a 3' to 5' exonuclease activity.

Phenylketonuria (PKU)

every newborn is tested: looking for buildiup of phenylalanine (amino acid): if it's located they have PKU. must go on special diet. caused by change in just ONE amino acid phenylalanine is not allowed to build up in wild type: its turned into tyrosine but buildup of phenylalanine can be toxic for people with PKU

Huntington's Disease, Hemophilia, Fragile X Syndrome are examples of the consequenes of what type of mutation?

examples of insertion mutations (repeated or inserted sequences of DNA)

How does base substitution become permanent?

first generation: replication error second generation: mutated molecule because parent is always right: adds complementary base, doesn't go back and correct error.

gene- and gene+

gene- : mutated gene (lacI -) gene+/gene : wild type gene. the plus sign can be left out. (lacI+)

Insertion/deletion mutations

large scale DNA insertions and deletions considered chromosome level mutations deletion: usually results in null phenotype insertion: results in null phenotype if in coding region, caused by TRANSPOSONS or repetitive sequences

How is a gene name typically written?

name in italics and lowercase first letter

Multi cellular organism somatic mutation: what type of cell is a somatic cell? Which cells are the mutations passed to? how will this affect survivability of species? example of this?

non sex cell: passed to daughter cells in the area. won't affect survivability of the species but can highly affect individual ex: individual exposed to too much ultraviolet light and they get a tumor but you don't pass on cells in skin to your offspring.

What would be the effect of an in-frame deletion of 3 bases? what is this mutation categorized as?

the loss of an amino acid. the rest of the protein is unaffected. this isn't really a frame shift because it didn't change the reading frame. still can have large affect on amino acid, though very specific event that is thrown into this category because no where else to put it

Which position in the nucleotide is usually changed in same sense mutations?

third position

DNA polymerase III's job in repairing a mismatch

to pick up at the cleavage site. And remember, the cleavage site, because it's the point of a nuclease activity, would be presenting a free 3' hydroxyl, so you don't need to prime this, you just bring DNA polymerase in, it adds in those bases, and presumably won't make the same mistake again. Even if it did, it's probably going to correct it via proofreading.

Identify if the following statement is True or False. Once DNA is fully methylated, the mismatch repair enzymes have no way of distinguishing between the parent and daughter strands.

true