Quiz 4 Flashcards
Purines
Adenine
Guanosine
Xanthine
Hypoxanthine
Pyrimidines
Cytosine
Uracil
Thymine
Uracil nucleoside and nucleotide
Uracil - Uridine - Uridylate
Cytosine nucleoside and nucleotide
Cytosine - Cytidine - Cytidylate
Hypoxanthine nucleoside and nucleotide
Hypoxanthine - Inosine - Inosinate
Xanthine nucleoside and nucleotide
Xanthine - Xanthosine - Xanthylate
PRPP Synthetase inhibitors
IMP, AMP, GMP, ADP,GDP
PRPP Aminotransferase inhibitors
IMP, AMP, GMP, ADP, GDP, ATP, GTP
Adenylosuccinate synthetase inhibitor
AMP
IMP Dehydrogenase inhibitor
GMP
Purine salvage pathways
Hypoxanthine/Guanine to IMP,GMP
Adenine to AMP
Carbamoyl-P Synthetase II Inhibitor
UMP, UDP, UTP, CTP
Carbamoyl-P Synthetase II Activator
ATP
Aspartate Transcarbamoylase Inhibitor
CTP
Aspartate Transcarbamoylase Activator
ATP
Difference between CPS1 and CPS2
1 is in the mitochondria and for the urea cycle, 2 is in the cytosol
Nucleoside Diphosphate Kinase specificity
Can be used for any of the diphosphate to produce CTP
2 sites of ribonucleotide reductase regulation
Site 1: dATP binding inhibits enzyme
Site 2: binding of triphosphate triggers reduction to diphosphates
dATP or ATP binding to site 2 is reduced to
UDP, GDP
dTTP binding to site 2 is reduced to
GDP
reduction of UDP, CDP is inhibited
dGTP
ADP
Thymidylate synthase inhibitor
F-dUMP
- made from fluoruracil, fluorodeoxyuridine
Dihydrofolate reductase inhibitor
Aminopterin
Amethopterin (methotrexate)
- these are folate analogs
Why is dTMP a good target for chemo
It is only used for DNA
Allopurinol
Inhibits Xanthine Oxidase
What causes gout
Too much purine synthesis leading to accumulation of uric acid
- deposits in joints/kidney
- caused by foods high in nucleic acids, proteins, alcohol, diuretics
Lesch-Nyhan syndrome
Absence of HGPRT
- symptoms: gout, self-mutilation, intellectual disability
3 ways chemotherapy works on cancer
1) blocks nucleotide biosynthesis
2) kills proliferating cells
3) blocks DNA replication (nucleotide analogues)
Fluorouracil, Fluorodeoxyuridine
Creates F-dUMP which works as a nucleotide biosynthesis inhibitor
Methotrexate
Folate antagonist, kills proliferating cells
AraC
Arabinose analog of deoxycytosine
- blocks DNA replication with nucleotide analogues
Acyclovir, Gancylovir, AZT
Chain terminators
- acyclovir, gancyclovir for Herpes
- AZT for AIDS
5-iododeoxyuridine, araA, araC
Chain elongation inhibitors
- 5-iodo for Herpes
- araA for viral encephalitis
- araA is a more potent inhibitor of viral polymerase vs host polymerase
B form vs A form hydration
B from is hydrated, while A form is present in low humidity
- changes the shape of the helix
Z form DNA significance
GCGCGCGCGC
- spreads out genes being actively transcribed
Secondary DNA stabilizing factors
H bonds, vDw, ions
Secondary DNA destabilizing factors
electrostatic repulsion, phosphate at pH 7 (negatively charged)
Why does ssDNA absorb more light
Absorbs more light (260 nm) bc the bases are exposed
Tm
50% of DNA denatured
5 types of drugs included in HAART
1) Nucleoside reverse transcriptase inhibitors
2) Non-nucleoside RTI (NRTIs)
3) Protease Inhibitors
4) Entry Inhibitors
5) HIV Integrase Inhibitors
Non-nucleoside reverse transcriptase inhibitors (NRTIs)
Inhibit addition of nucleotides to chains using a nucleoside analog
- acyclovir, AZT
Negative supercoil
Underwound
Positive supercoil
Overwound
Is natural DNA negative or positive
Negatively supercoiled
Topoisomerase 1
break one strand, change by +1 or -1 supercoils
- requires DNA to be strained + or -
- does not affect unstrained DNA
Topoisomerase 2
cuts 2 strands, changes number of supercoils by 2
- can add supercoils to new DNA in prokaryotes only
How do topoisomerase inhibitors work
Inhibit re-ligation function by the enzyme
Topoisomerases 1/2 in prokaryotes and eukaryotes
Different version in each
Histones
small, positive proteins
H1 function
Linker DNA, higher-order structures
H2A, H2B, H3, H4 function
Nucleosome core
- DNA wrapped around is same length as Okazaki
Solenoid
35-40x shortening of DNA
- 6 nucleosomes per turn
Highly repetitive sequences
- short, tandem, AT-rich
- 10% of genome
- telomeres
- greater than 300,000 copies per genome
Moderately repetitive sequences
- code for highly-used genes
- SINES/LINES space out actively transcribed genes
Unique or single-copy sequences
- only 1% made (housekeeping genes)
- grouped in families
- pseudogenes: non-functional unique sequences
Myotonic Dystrophy Type 1
- more CTG repeats increases severity (onset age decrease)
- due to polymerase slipping on repeats
What is anticipation
increase in severity of a disease with each successive generation
Origin of replication
P - OriC
E - ARS
Protein that binds origin
P - dnaA complex
E - ORC
Helicase protein
P - dnaB/dnaC complex
E - MCM
Protein that stabilizes ssDNA
P - SSB
E - RPA
Protein that increases processivity
P - ß-subunit polymerase 3
E - PCNA
Clamp loader
P - gamma complex
E - RFC (replication factor C)
Protein that makes RNA primer
P - dnaG (part of primosome)
E - DNA pol ⍺
Protein that removes RNA primers
P - DNA Pol 1
E - RNAse H and Fen1
How do dnaA and ORC function?
Bind DNA binding domain and using positive histone-like proteins, causing DNA to split apart at nearby 3 AT-rich regions
How does priming work in prokaryotic replication
Provides an uncreated 3’ OH, allowing DNA polymerase to bind
- RNA primer is 5-10 nts
DNA Pol I
Primer excision and DNA repair
- 3’ exonuclease: proofreading, removes wrong brase
- 5’ exonuclease: removes primers (nick translation)
Nick translation
5’ exonuclease removes one base at a time until it gets to DNA
- DNA ligase then seals the open gap
DNA Pol III
bulk of replication, high processivity and rate
Core enzyme of DNA Pol III
Dimer of ⍺, ε, θ
ß-sliding clamp
Improves processivity, contains negative amino acids on interior to help DNA slide through
Function of χ subunit on 𝛾 complex
Helps transition DNA Pol III from RNA primer to making DNA
Replisome
Creation of a loop at at replication fork so DNA Pol III goes in the same direction on the lagging and leading strand
Proofreading in prokaryotes
epsilon subunit removes mismatched base with 3’ to 5’ exonuclease
Termination in prokaryotes
- ter sites: x6, 3 each side apposed 180º from OriC
- TUS protein: binds ter sites in opposite direction
- Type II topoisomerase: separates new interlinking strands, then religates
Protein that synthesizes DNA
P - DNA Pol III
E - DNA Pol ε, ẟ, 𝛾
DNA Pol 𝛾
Only in mitochondria
DNA Pol ε
Leading strand replication
DNA Pol ẟ
Lagging strand replication
- High processivity with PCNA, low without
DNA Pol ε
Leading strand replication
- High processivity regardless of PCNA
Licensing
Assembly of pre-replication complex on ARS in eukaryotes
- ORC, inactive MCM, Cdc6, Cdt1
Firing
Helicase activation, which activates complex to begin replication
hTERT
Protein component of Telomerase
- reverse transcriptase
hTR
Template for hTERT (RNA component)
t-loop
Leftover overhang that comes in and displaces ds-DNA to make D-loop-t-loop
Dyskeratosis Congenita
Mutations in hTERT/hTR cause premature mortality
Normal methylation in prokaryotes
Adenine and Cytosine
Normal methylation in eukaryotes
Only Cystosine
Methylation Heritability
1) Sites chosen during gametogenesis and embyrogenesis
2) not all C’s methylated
3) maintenance methylase after replication
How does methylation control gene expression
Un-methylated promoter: expressed
Methylated promoter: not expressed
5-azacytidine
N instead of CH3 prevents methylation
- allows gene expression
Transition mutation
Purine-Pyrimidine to Purine-Pyrimidine
- GC -> AT
Transversion mutation
Purine-Pyrimidine to Pyrimidine-Purine
- AT -> TA
- can’t be fixed
Photodimerization
Dimerization of adjacent intra-strand pyrimidines by UV
- creates Thymine dimers
8-oxoguanine mispaired with Adenine
Caused by ROS
Types of single-strand repair
1) direct base repair
2) excision repair
3) mismatch repair
Mechanism of direct repair of base in single-stranded repair
MGMT transfers methyl group onto itself from O6-alkylguanine
- alkylates itself and restores Guanine
Mechanism of NER in prokaryotes
1) UvrAB recognizes damage and bends DNA
2) UvrA leaves, UvrC joins
3) 3’ cut, 5’ cut, helices removes damaged piece
4) DNA Pol I replaces excised DNA, ligase seals nick
Mechanism of base excision repair in prokaryotes
1) Uracil-DNA n-glycosylase recognizes and removes damaged base from backbone
2) Endonuclease creates 5’ nick
3) Backbone removed and replaced with new base by DNA Pol I 5’ endonuclease
