Molecular Biology Flashcards
Nucleotide
- sugar
- base
- phosphate
deoxyribose
- DNA
- missing 2’ OH
- less apt to nucleophilic attack
nucleic acid polymerization
- 5’ to 3’ synthesis and base sequence
- antiparallel and complementary
- phosphodiester bond
phosphodiester bond
- covalently links nucleotides between 3’ OH and 5’ Phosphate
bases
- purines
- pyrimidines
purines
- 2 rings
- adenine
- guanine
pyrimidines
- 1 ring
- cytosine
- thymine
- uracil
genome
- all the DNA in an organisms
prokaryotes
- single circular DNA genome
- methylation
- supercoiling
methylation
- protection from their own restriction enzymes (endonucleases)
- no longer fits into the active site of the enzyme
endonucleases
- chop off DNA
- restrict the growth of viruses whose DNA is not methylated
supercoiling done by the enzyme
- gyrase - uses ATP by breaking the DNA and twisting the two sides of the circle into supercoils.
eukaryotes
- several linear chromosomes
packing of eukaryotes
- DNA packaged with histone octomer to form nucleosomes
- forms the bead on a string
- packed further into chromatin
- packed into the eukaryotic cell as a chromosome
heterochromatin
- tight packing inactive DNA
- dense, dark regions on a stain
- rich in repeats
euchromatin
- less tightly packaged
- active DNA
- higher transcription rates and gene activity because DNA more accessible to enzymes and proteins
histones
- basic to attract acidic DNA backbone
- two of each: H2A, H2B, H3, H4
centromere
- region on the chromosome where
- mitotic spindle attaches via kinetochores during cell division
- where two pieces of DNA are held together after replication
centromere position
- position defines ratio between long and short arms
equal size arms
- metacentric
really short P arms
- acrocentric
no P arm
- telocentric
short P arms
- submetacentric
telomeres
- ends of linear chromosomes
- “caps” linear chromosomes to prevent degradation
- many repeats of a short sequence
ends of linear chromosomes
- loops of ssDNA to protect ends of chromosomes
telomere caps
- prevents activation of repair pathways
- prevent fusion with neighboring chromosomes
do prokaryotes have telomeres?
- NO!
- they only have a circular chromosome
start codon
- AUG - methionine
stop codons
- UAA - U Are Annoying
- UGA - U Go Away
- UAG - U Are Gone
- specify no amino acids
degenerate
- multiple codons for the same amino aicd
intergenic regions
- never transcribed nor translated
- no genes
- inherit the same intergenic regions from our parents.
polymerase errors
- point mutations
- small repeats - DNA pol falls off DNA strand then rejoins.
- insertions/deletions (small, frameshift)
endogenous damage types
- inside the cell
- reactive oxygen species
- physical damage
endogenous damage effect on DNA
- inside the cell
- oxidized DNA - bases look different so they can’t base pair.
- crosslinked bases - physically linked together. not just hydrogen bonded. can’t separate strand easily for replication. could crosslink to different strands.
- physical damage - DNA broken or bases missing. sheer stresses.
- these can lead to polymerase errors
exogenous damage types
- outside of the cell
- radiation
- chemicals
exogenous damage effect on DNA
- UV rad - pyrimidine dimers. pair with each other. (typically T=T)
- X rays or gamma rays - double stranded breaks and translocations
- chemicals - can lead to physical damage or to intercalation and polymerase will stick in something random and cause errors.
transposons
- insertions/deletions
- inversions
- duplications
point mutations
- single base pair change
missense mutations
- codon for aa becomes a new codon for new aa
- changes aa
nonsense mutation
- codon for aa becomes STOP codon
- shortened protein
silent mutation
- codon for aa becomes new codon for same aa
- no effect
frameshift mutations
- insertions and deletions
- changes the reading frame
transposase
- cut and paste enzyme
- allows for mobility
IS element (inverted sequence)
- transposase only
Complex transposon
- transposase with genes
composite transposon
- two transposase flanking a central region
how transposons contribute to genomic variation
- code for the cut and paste transposase enzyme
- transposase cuts the transposon out
- transposase pastes transposon somewhere else.
if a transposon is inserted into the intergenic region
- no effect
if a transposon is inserted into the coding region
- can result in a big mutagenic effect
if two transposons are in the same direction
- pair up
- big deletions and possible insertions
- chromosomal rearrangements, possibly on a different chromsome
if two transposons are in different directions
- pair up to form U-loop
- flipped and lead to inversion
- chromosomal rearrangement on same chromosome
inversion
- a reversal of the gene sequence
amplifications
- doubling of the gene
mismatch repair pathway
- repairs bases due to DNA polymerase errors
- detected after replication is complete
- fix DNA based on polymerase errors
- methylate the parent strand
- cut out incorrect bases that are unmethylated
- polymerize again
base/nucleotide excision repair
- occurs prior to replication because defective bases will lead to polymerase errors
- incorrect base excised and replaced
homologous end joining
- repairs double stranded DNA breaks
- occurs after replication
- sister chromatid used as a template for repair broken strand
- crossover of the sister chromatid
- may lose an allele
- best way to repair double stranded breaks
non-homologous end joining
- no sister chromatid for template (cell is not going through division and thus not replicating DNA).
- clear out damaged regions
- blunt ends of DNA
- join the broken 2 strands together
- mutagenic because losing some bases or could result in a translocation
translocations
- due to recombination between non-homologous chromosomes or faulty DNA repair (non-homologous end joining)
- causes gene fusion if the joining point is in the middle of the gene.
direct reversal
- white light reverses damage
- pop pyrimidine dimers back into place to fix DNA
rules for carrying out DNA replication
- semiconservative - half of original DNA molecule will be saved in new DNA molecule
- 5’ to 3’
- requires an RNA primer
- requires a template
helicase
- unwinds DNA
topoisomerase
- cuts DNA
- relaxes supercoiling by passing strands through each other.
primase
- puts down the RNA primer
DNA polymerase
- replicates DNA, proofreads, removes primer
ligase
- links Okazaki fragments
prokaryotic replication
- theta replication
- 1 origin
- 5 DNA polymerases
DNA pol III
- high processivity
- fast 5’ to 3’ polymerase and 3’ to 5’ exonuclease activity
- main replicating enzyme
- no known function in DNA repair
DNA pol I
- low processivity
- adds nucleotides to RNA primer then DNA pol III takes over
- slow 5’ to 3’ polymerase and 3’ to 5’ exonuclease
- 5’ to 3’ exonuclease to remove primer
- DNA excision repair
DNA pol II
- 5’ to 3’ polymerase and 3’ to 5’ exonuclease
- back up for DNA pol III
- DNA repair
DNA pol IV and V
- error prone 5’ to 3’ polymerase activity
- DNA repair
eukaryotic replication
- multiple origins of replication
- replication bubbles
- several DNA pols, complex multisubunit enzymes
end replication problem
- primers add at lagging strand
- RNA primers removed
- shorter telomeres
telomerase
- lengthens the telomeres by adding bases
telomerase characteristics
- carries his own internal RNA primer to lengthen telomeres
- reverse transcriptase activity
hnRNA
- heterogenous nuclear RNA
- unprocessed RNA in euk
- precursor to mRNA
miRNA
- microRNA
- helps regulate gene expression
siRNA
- small interfering RNA
- used for regulating gene expression
- forms ds RNA molecule helix and ribosome will fall off.
coding strand
- sense strand
- same code but T instead of U
template strand
- anti-sense strand
- complementary
- the strand being transcribed
regulation of transcription
- promoter - binding site for RNA polymerase
strong promoter
- high affinity for RNA pol
- get a a lot of RNA
- high rates of transcription
weak promoter
- low affinity for RNA pol
- low rates of transcription
DNA binding proteins
- repressors - gene on must be turned off. bind to DNA to stop transcription.
- enhancers - gene off must be turned on. Bind to promoter.
prokaryotic transcription
- transcription and translation happen in the cytosol at the same time
- no mRNA processing
- polycistronic - code for several different proteins from same mRNA
- 1 RNA polymerase
eukaryotic transcription
- transcription (nucleus) and translation (cytosol) in different places
- mRNA processing
- monocistronic - one mRNA one protein
- 3 RNA polymerases
mRNA processing
- 5’ G cap
- 3’ poly A tail
- splicing - remove introns. keep exons.
RNA polymerases
- RNA pol I - rRNA
- RNA pol II - mRNA
- RNA Pol III - tRNA
are empty - rmt
aminoacyl tRNA synthetase
- attaches AA to correct tRNA
- individual for all codoons
wobble hypothesis
- first two anticodons on tRNA bind normally
- third anticodon is more loosely bound
- adenine on tRNA can get converted to idenosine
- happens when there are G, U, I at 5’ end of anticodon
prokaryote ribosome subunit
eukaryote ribosome subunit
- 50S + 30S = 70S
- 40S + 60S = 80S
P site
- growing protein held here
- first tRNA binds here
A site
- new amino acid added here
initiation of translation
- ribosome subunits assemble over the mRNA with met and tRNA in the P site
elongation of translation
- add new AA in A site
- costs 1 GTP
- form peptide bond between 2 AA
- ribosome moves forward one codon
termination of translation
- stop codon in A site
- bind release factors
- break bond between final tRNA and final AA releasing the completed protein
tRNA loading costs how many ATP
- 2 ATP per tRNA
initiation costs how many ATP
- 1 ATP
A site binding costs how many ATP
- 1 ATP per tRNA
- 1 less than number of AA
translocation costs how many ATP
- 1 ATP each time
- 1 less than number of AA
termination costs how many ATP
- 1 ATP
how to calculate the number of ATP needed
amino acids x 4
post translational modifications
- protein folding
- covalent modification
- processing
protein folding
- helped by chaperones
- H bonds
- hydrophobic/philic interactions
covalent modification
- disulfide bridges
- glycosylation
- phosphorylation
- methylation
processing
- remove some parts of a protein to make it active
- zymogens
zymogens
- inactive enzymes
- pro/ - ogen
areas of the DNA that are easily unwound for replication are composed of what bases?
-A-T rich
stop site
- usually a polyadenylation signal
- different in prokaryotes
operator region
- where repressors bind
repressor
- prevents RNA polymerase from binding to protein
amino acyl tRNA synthase
- attaches amino acids to correct tRNA.
- need as many unique ones as we have unique tRNAs.
