Nucleic Acids and DNA Synthesis Flashcards
Nitrogenous bases
Purines- adenine, guanine
Pyrimidines- cytosine, thymine
Aromaticity of pyrimidines/purines
Can convert between the lactam and lactim forms, lactam (keto) form dominates at physiological pH
N-glycosidic bond
Bond between the base and the sugar, normally in the anti conformation
Nucleoside vs. nucleotide
Nucleoside- base and sugar
Nucleotide- includes the base, sugar, and phosphate(s)
Phosphodiester bond
Linkage between two nucleotides
Sugar-phosphate backbone
Multiple phosphodiester bonds that make up the nucleic acid
5’ end
End of the polymer with phosphate attached to C5’
3’ end
End that has a free OH group at C3’
Base pairing
DNA contains two polynucleotide strands that base pair through hydrogen bonds, each contains one purine and one pyrimidine
Chargaff’s rules
[A] = [T], [G] = [C], purines = pyrimidines
Double helix
Two antiparallel strands base pair together are twisted to form double helix
Z-DNA
Transient form in cells- line connecting phosphate zags
B-DNA
Dominates in living systems
A-DNA
Dominates in DNA-RNA hybrids, tightly wrapped, dehydrated form of B-DNA
Structure of RNA
RNA has many forms, structure related to function, typically single stranded, has secondary structure, similar to DNA structure
Ribosomes
Site of protein synthesis in the cell, contains large and small subunits, comprised of protein and RNA, rRNA is important to ribosome function
Prokaryotic ribosomes
Total size 70S- 50S + 30S, three types of rRNA, 23, 16, 5S
Eukaryotic ribosomes
Total size 80S- 60S + 40S, four types of rRNA, 28, 18, 5.8, and 5S, mitochondrial ribosomes- 55S, similar to prokaryotes
tRNA
tRNA secondary structure is important in protein synthesis, anticodon loop is responsible for recognizing the codon on the mRNA, tRNA bring in the amino acid being added to the polypeptide
Anticodon
Three base sequence that base pairs with mRNA
Denaturation
Separation of the double helix into two separate strands, heat can disrupt hydrogen bonds and cause DNA denature, basic solutions denature DNA and RNA
Tm
Temperature where 50% of DNA is denatured, determined by the amount of G:C vs. A:T, changed by salt concentration (stabilizes the solution)
Basic solutions
Denatures DNA, causes RNA to break apart because the OH group on ribose loses its proton, negative charged oxygen breaks phosphodiester bond
DNA hybridization
DNA can be hybridized to a complementary strand of RNA, temperature reduced slowly
Packing of DNA
Supercoiled, bound to histones to form chromatin
Histones
Major protein that forms chromatin, high percentage of lysine and arginine (positively charged), H2A, H2B, H3, and H4 form nucleosome, H1 joins nucleosomes together
Solenoid structure
Helical structure of several nucleosomes
Human genome
Humans have 23 pairs of chromosomes (22 somatic, 1 sex), genes are arranged across each chromosome
Proteins involved in prokaryotic replication
Helicase, topoisomerase, gyrase, single-strand binding proteins, DNA polymerase (I, II, III), primase, DNA ligase
oriC
Sequence in the prokaryotic genome where replication is initiated- DnaA binds and helix unwraps, two replication forks
Semiconservative replication
After replication, one new strand and one parental strand
Prokaryotic replication- unwinding the strands
Helicase unwinds the DNA strands, topoisomerase breaks/rejoins phosphodiester bonds to relieve supercoiling, single-strand binding proteins bind to DNA as it unwinds to prevent re-associating and enzyme degradation
Prokaryotic replication- DNA strands are primed
Primase adds RNA primers to DNA (DNA polymerase needs 3’ OH), after DNA is synthesized the primers are removed by RNase H and DNA pol I
Prokaryotic replication- new strands synthesized
DNA polymerase III adds the next deoxyribonucleotide making a new strand 5’ to 3’, proof reads as it functions, produces leading and lagging strands
Leading strand
Synthesized continuously 5’ to 3’ toward replication fork
Lagging strand
Synthesized discontinuously 5’ to 3’ away from the replication fork, produces Okazaki fragments that are joined by DNA ligase
Differences between eukaryotes and prokaryotes
Eukaryotes have larger genomes, histones/nucleosomes, prokaryotic circular DNA
Eukaryotic origins of replication
Have multiple origins of replication, each origin has two replication forks, fork from one origin will meet replication fork from another, multiple origins makes it possible to copy complex genome
Prokaryotic DNA polymerases
Polymerase I- replication, repair, primer excision
Polymerase II- DNA repair
Polymerase III- major replication polymerase, 3’-5’ exonuclease for proofreading
Eukaryotic DNA polymerases
Polymerase alpha- associated with primase
Polymerase beta- DNA repair, primer excision
Polymerase gamma- mitochondrial DNA synthesis
Polymerase delta- replication, 3’-5’ exonuclease for proofreading
Polymerase epsilon- replication, 3’-5’ exonuclease for proofreading, DNA repair
Replication fork
Polymerase epsilon- replication of leading strand
Polymerase delta- replication of lagging strand
Problems with linear DNA
DNA polymerase cannot copy the end of the lagging strand, produces a 3’ overhang, telomerase adds nucleotides to the 3’ end but still have overhang
DNA methylation
DNA can be methylated after replication, adenine and cytosine, species specific, methyl groups project into groove and are bound by DNA binding proteins
Damage to DNA
DNA is exposed to chemical and physical agents that can damage it daily- smoking (benzopyrene), sun exposure (thymine dimer), x-ray (hydroxyl radical), deamination, translation-coupled repair
Base excision repair (BER)
Removal of a damaged base that cannot be directly repaired, glycosylase cleaves glycosidic bond, deoxyribose is cleaved by an AP endonuclease, residues are removed by an exonuclease, DNA polymerase fills the gap, DNA ligase seals the nicks
Nucleotide excision repair (NER)
Corrects large segments such as pyrimidine dimer or lesions with bulky substituents, endonuclease cleaves distorted regions and removes DNA segment, DNA polymerase fills the gap and DNA ligase seals the nicks
Mismatched repair
Function to fix errors in replication that are missed by proofreading, methyltransferase has not had time to methylate newly synthesized strand, determines which base is an error, steps similar to BER and NER
Homologous recombination
Crossing over of genes between homologous chromosomes
Translocation
Caused by breaks in two nonhomologous chromosomes, balanced translocation results in no loss of genetic function because the cut occurs between genes, unbalanced results in extra or missing genes
Transposons
Sequences of DNA that are moved from place to place in the genome, end of the transposon has inverted repeats that act as target sequence, staggered cuts produce single stranded ends that are ligated to the transposon
Reverse transcriptase
Makes DNA from RNA, known as cDNA, can be incorporated into the human genome, causes disease
Deamination
Cytosine loses the amino group and becomes uracil, can become thymine if methylated, disrupts base pairing and can permanently change DNA sequence, if DNA had uracil than deamination would be highly mutagenic
Equation for Tm
Tm (deg C) = 69.3 + 0.41 (%G + C)
