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111 Cards in this Set

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Proteins that are implicated in stabilizingn chromosome structure?
SMC proteins
What is the name of the bond that connects the sugar and the base in a nucleoside?
N-glycosidic bond
A left handed double helix is the ---- form
z
transcriptional regulators recruit
CRC - Chromatin Remodeling Complexes
CMC -Chromatin Modifying complexes
What are the two types of heterochromatin?
1) constitutative - these genes are silenced most of the time
2) facultative - normally present within euchromatin and is implicated in silencing
Which is ATP dependent -

CRC or CMC (chromatin remodeling complex or Chromatin modifying complex)?
Chromatin remodeling complexes
What is the role of remodeling complexes?
-dependent on energy
-dissociate the nucleosome and give access to other molecules to the DNA or to the Histone
-can change the position of the nucleosome relative to the DNA
-can change histones
What is the role of Chromatin modifying complex?
- generally modification of histone N terminal or C-terminal
-can be acetylators (HAT) or deacetylators (deactylase transferases)
-also involeved in methylation (methyl transferase)
Tell me about the disease ICF syndrome
- a disease in which DNA methylation process is impaired due to a mutation in methyltransferase
-the heterochromatin is not stable
-disease associated with immunodeficiency and facial abnormalitieis
allele
a form of a gene
The patient with the disease is called the
proband
Three ways that autosomal dominant can actually manifest itself
1) Haploinsufficiency - you need two normal functioning genes to make enough protein, and you only have one functioning copy, so you don't make enough protein
2) dominant negative effect - a mutatnt protein interferes withthe normal function from the other allele
3) two-hit suppressor model - you have one good gene and one bad gene, the good gene is knocked out and then the bad gene mutates.
Examples of autosomal dominant genes
1) Achondroplasia
2) FAP - colon polyps
Factors that can change the manifestations of autosomal dominant diseases
1) non-penetrance
2) delayed onset
3) anticipation
4) germ-line mosaicism
5) phenocopy effect
6) non-paternity
non-penetrance
the parent has a copy of the mutation, but doesn't show any signs of it - i.e. males can carry the breast cancer gene just as well as females, but they don't manifest the disease
2) delayed onset
-hunnington disease for instance may manifest itself in the late 30s after the proband has already reproduced
3) anticipation
disease gets worse from generation to generation - generally
4) germ-line mosaicism
mutation is found only in gametes, not in the stomatic cells -skin/blood of the parents
5) phenocopy effect
there are too many possible genes that could be causing the disease, for instance breast cancer, so you don't know which gene is responsible
clinical heterogeneity
same gene, different diseases; Ret for instance can either cause endocrine cancer or can cause hershsprung's
allelic heterogenity
different mutations on the same gene, lead to the same disease - cystic fibrosis for example

in a population there may be a number of different mutant alleles at a single locus. The same or a similar phenotype may be caused by different mutant alleles rather than by identical alleles at the locus.
locus heterogenity
different mutations in different genes, same disease -hearing genes, colorblindness

the production of identical phenotypes by mutations at two or more different loci
variable expressivity
same defect, but different manifestations of the SAME disease. for instance one child could be bed ridden and the other one could be running around.
example: cafe au lait spots
Examples of anticipation
huntington's
fragile x
myotonic dystrophy
What is the probability that an affected male with CF will have an affected offspring
zero. Males with CF are generally sterile
Daughters of affected males are ---- (hemophelia for instance)
Obligate carriers if the disease is an X-linked disease
Sons of affected males are --- (hemophelia for instance)
normal. Males are never affected when their father caries the x-linked diseases (mother must be normal)
What is the risk of a carrier mother to have a son with the disease (hemophelia)
1/4, 50% chance of having a boy, 50% chance of having an affected child.
what is heteroplasmy?
In mitochondrial related diseases - becasue mitochondria segregates randomly, you can have different numbers of mitochrondria (and diff numbers of mutated mito) in different cells

Disease is associated with degree of heteroplasmy.
which structure (level) in amino acids can accomodate ligand binding
quaternary strucutre
-for instance, the hemoglobin subunits have to associate to a quaternary structure for them to be fully functional and to be able to respond to changes in oxygen tension
amino acids in proteins that are derived from mRNA are almost always which type of isomer?
- levorotary (the L-isomer)
Lys
lysine
basic
+ charged
soluble in water - on the outside of a protein
Arg
arginine
basic side chain
+ charged
soluble with water
His
Histidine
basic side chain
+ charged
soluble with water
Asp
Aspartic acid
acidic side chains
-charge
soluble with water
Glu
glutamic acid
acidic side chain
- charge
soluble with water
Asn
Asparagine
polar side chain
soluble with water
Gln
glutamine
polar side chain
soluble with water
Ser
Serine
polar side chain
soluble with water
Thr
Threonine
polar side chain
soluble with water
Tyr
Tyrosine
polar side chain
soluble with water
This aa can be found at the border of the hydrophobic and hydrophilic?
Tyrosine

potentially also Thr
Ala
alanine
nonpolar side chain
Val
valine
nonpolar side chains
Leu
leucine
nonpolar side chain
Ile
isoleucine
nonpolar side chain
Pro
proline
nonpolar side chain
Phe
phenylalanine
nonpolar side chain
Met
Methionine
nonpolar side chains
Trp
Tryptophan
nonpolar side chain
Gly
glycine
nonpolar side chain
Cys
cysteine
nonpolar side chains
What two aa's have two chiral centers?
Thr
Ile
This aa is not really an amino acid, but is an imino acid
proline
not commonly found in secondary and tertiary structures becasue it introduces some space in the peptide bond
has an imido group
asparagine
glutamine
has an indol group
tryptophan
nonpolar side chains contain ---
aliphatic groups
like alanine, valine, leucine, isoleucine, proline, phenylalanine.
a classic example of modified amino acids
4-hydroxyproline + 5-hydroxylysine
(basically a hydroxyl group has been added to a proline and a lysine)
-this is found in collagen.

Other examples:
gamma-carboxyglutamate : carboxy group added onto glutamate

histones are methylated on their lysines
which aa's can form salt bridges
negatively and positively charged aas:
lys, arg, his, asp, glu
what do we need to know about titration curves?
- you can use titration curves to determine the net charge of the amino acid
I have an aa, i raise the pH, what groups will loose protons in what order?
first the carboxylic function will loose its proton, (then the carboxylic group on the side chain if it exists), then the amino group will looses its proton
list non-standard amino acids with hydroxy modifications
4-hydroxyproline + 5-hydroxylysine
list non-standard amino acids with phosphorylation modifications
tyrosine
threonine
serine can all be phosphorylated
two most important modifications to aa groups
hydroxylation (proline + lysine)
phosphorylation (thr, tyr, ser)
Are small or long peptides generally biologically active? e.g.?
small peptides are biologically active
example glutathione.
what can we get from the titration curve
-titration curves give an estimation of the pkas of each ionizing group and theri corresponding buffering zone

-titration curves help predict the electric charge of the amino acid (at a given pH)
What helps open up the double helix for DNA replication?
Initiator proteins
What's the difference between mammalian and bacterial DNA replication in terms of replication of origin?
-bacterial - only have one origin of replication at a time

-mammalian - have multiple origins of replciation, and they are all bi-directional (this makes up for the fact that mammalian DNA replication is much slower than bacterial DNA replication)
What makes DNA replicaton more efficient?
- the association of the leading and lagging strand polymerases III together

- the sliding clamp
Which mammalian polymerase types are associated with DNA replication?
polymerase alpha and delta are associated with DNA replication
Difference between Topo I and Topo II?
Topo I cuts one strand DNA
(involved in DNA synthesis during the S phase)

Topo II cutes double stranded DNA
(involved in mitosis)
Which topo is involved in DNA synthesis?
TOPO I

(Topo II is involved in mitosis)
3 differences between bacterial replication and mammalian replication?
- bacterial - 10X faster replication, doesn't have histones, only one origin of replication

-mammalian - slow replication, yes, has histones, has multiple origins of replication
What is special about insulin in terms of tertiary structure ?
Insulin is stabilized by three disulfide bonds that are formed from the oxidation fo cysteine residues on the two polypeptides - this is unusual because it is a COVALENT bond
conservative substitution
when a modified protein is completely intact interms of function despite the mutation

(non-conservative substitution is when the enzyme function is changed by the mutation )
Describe the polypeptide structure
polypeptides are basically a series of planes that rotate around each other

the peptide bonds are uncharged

NC bond is the phi angle
CC bond is the psi angle
Where are beta turns typically found?
At the surface of the protein, so that they can interact with the environment and stabilize the protein
For Hardy Weinberg calculations if you get a q^2 of 1/100000 or greater, what can you assume for your calculation
As q approaches zero, p goes to one, so the frequency of heterozygotes is 2q.q
what's special about x linked diseases and affected males?
The incidence of the disease is equal to the frequency of the allele if you are looking at affected males, because males are hemizygous.
coefficient relationships
what genes are in common
basically you count the number of branches and then multiply by 1/2 for each branch ... each branch gives you a half contribution.

i.e. you have fist cousins and you are looking at the risk that they would pass a gene on the chance that they inherited that gene from a previous relative up top would have been 1/8.
Things that screw up H-W
- assortative mating (stratification)
-selection & fitness
-genetic drift, gene flow, founder effect, consanguity,
How do you calculate the degress of selectivity?
degree of selectivity = 1 - fitness.

For instance, fitness for achrondroplasia is 1/5 or 0.2
so the degree of selectiity is
1-0.2 = 0.8. This is also equivalent to the % of new mutations ...
What does fitness equal (equation)
Fitness = 1-S (degress of selectivity)
How do you calculate the % of disease caused by a new mutation?
We look at the fitness. For instance if the rate of reproduction is 1/5 that of the normal population, the fitness is .2, and therefore 0.8 or 80% is the percentage of new mutations. (0.8 + 0.2 = 1) because 1 = total number of kids with disease, 0.2 is the amount of kids that genetically inherited the disease, and 0.8 is the number of kids whose gene randomly mutated.
If you have lethal fitness, what is the percentage new mutation?
100% because f = 0, so 1-0 = 1 or 100%
Polymorphisms
-variations in a gene that are very common
-polymorphisms usually don't cause a disease, but they can predispose you to a disease
-we have polymorphisms because they generate diversity
An example of a polymorphism predisposing people to a disease
B27 variatnt
ankylosing spondylitis - curvature of the spine
95% of people with the polymorphism have the disease
How do you know when a disease is multifactorial?
- genetic predisposition + environment = multifactorial inheritence
-risk for a disease is higher in the family than in the regular population
-BUT the disease doesn't look like recessive or dominant inheritance
-not dominant becasue incidence of the disease goes down by more than half as you go through the family
-not an autosomal recessive becasue more than 25% of sibs on average are effected
-twin studies are often used to prove multifactorial inheritance - contribution is genetic but a lot may be environment
examples of diseases that are multifactorial
-ankylosing spondylitis
-cleft pallet
-pyloric stenosis (projectile vomiting) - this is more common in boys than girls
Explain recurrence risks in multifactorial diseases
-generally the larger number of people in your family that have the disease, the higher your risk for getting it
-the more severe the form of the disease, the higher the risk
-if the frequency of the disease in the population is very small, and your family has the rare disease, then you have a higher risk
-if there are sex differences, and the proband is the less common sex.
how do restriction endonucleases recognize the clevage site, and also, how frequently do they make cuts?
- normally recognize a site with palandromic symmetry to it

-for a 6 base sequence - you should get once every 4,000 bases 1/(4^6)

- for a 4 base sequence - you shuold get once every 256 bases which is 1/(4^4)
name some restriction endonucleases (common)
-Bam
-EcoRI
-Sau3A
most cloning vectors are ----
plasmids
What are the three critical critera of a successful vector
-a site to insert the target DNA = called a POLYLINKER INSERTION SITE
- an origin of replication
-a selectable marker like amp resistance
what is the most common host for vectors? What is an important consideration to think about?
-E. cori is the most common host
-must think about post-translational processing/modification - this will be different in a bacteria vs. a eukaryote.
what enzyme is responsible for making a cDNA library?
reverse transcriptase - it can take the mRNA and make the first DNA strand from it, so that you have a DNA-RNA hybrid ...`
How do we identify clones?
DNA - southern blot
RNA - Northern Blot
Protein - western blot
What temperatures do you use for PCR?
94 degrees for denaturation
55 degrees for annealing
72 degrees for DNA extension
What elements are required for PCR
-oligonucleotide primers (no primase) - it just anneals
-heat stable taq polymerase
When PCR is used for forensics - what are they looking at?
- they look at different regions of the cell where the DNA is repeated over and over - these so called repetitive elements like CACA ... because some people have three copies of this and another guy might have ten copies of the same thing. If you amplify this DNA you will come up with different bands
fixative for light microscopy
formaldehyde
fixative for EM
gluteraldehyde
OsO4
LM tissue samples are embedded in
paraffin wax
EM tissue samples are embedded in
plastic - epoxy resin + acrylate resin
what dye, when excited can bind nucleic acids
DAPI
resolution in LM
0.2 um
resolution in EM
1.0 nm
LM embedded tissues can be sliced ...
5-15 um thick
EM embedded tissues can be sliced
50-150 nm thick
name two selective dyes and what they are used for
PAS - a stain used for carbohydrates

Toluidine - a blue metachromatic dye that can be used to class proteins
expalin
"acidiophilic"
and basophilic dyes
acidophilia - acid loving. EOSIN dye is an acid dye, it has a net negative charge and binds to net positive charges

basophilia - HEMATOXYLIN is a basic dye with a net + charge that binds - charges