Molecular Biology And Biochemistry DNA Flashcards
Characteristic components of nucleotides
Nitrogenous base
One or more phosphates
A Pentose sugar
Why is ribose sugar more reactive the deoxyribose
It’s more reactive and unstable because of the OH group on the 2 carbon
Which bases are purines
Adenine and guanine
Which bases are pyrimidines
Thymine, uracil and cytosine
What is a pyrimidine
Single ring nitrogenous base
Structure of dna
Two anti parallel strands
Each strand has a 5’ (phosphate) and 3’ end (hydroxyl)
Held together by hydrogen bonds
Antiparallel nature allows for h bond formation
Bases planar to sugar phosphate backbone
What evidence proved dna helical structure
X ray diffraction
How does the x ray diffraction of dna show a helical structure
X shape which is indicative
Layer line which shows the length of one complete turn
The common features of B-DNA
Right handed double helix
Hydrophobic core
Hydrophilic backbone
Major and minor groves
Factors towards the stability and structure of the double helix
Backbones separated as far apart as possible to avoid electrostatic repulsion
Bases form stablizing h bonds
Bases stack to maximise aromatic ring interactions
How many hydrogen bonds form between the bases
A +t 2
C + g 3 (
Why doesn’t non standard base pairing work
Distorts the geometry of the double helix the bases are no longer 1.1 nm apart
Why are their aromatic stack interactions
Benzene contains an e in all p orbitals which can delocalise and form a ring of electron density around the planar ring
Why is displacement stacking better then eclipse stacking
Maximises attractive interactions between dipoles
Why are there major and minor groves
The glycosidic bonds between bases are the same distance apart so nucleotides must be at an angle
They allow for dna bonding proteins to read dna without unwinding the helix
Why does DNA have thymine not uracil
Cytosine can delaminate to uracil therefore uracil is identified and removed in dna repair processes
Why does RNA have uracil not thymine
Thymine is more energetically costly to produce as a methylated uracil
There’s no bio synthetic pathway for it to be produced
Why is rna less stable then dna
OH group on c2 acts as an internal nucleophile and causes the breakage of the phosphodiester backbone
What end are nucleotides added to in replication
3’
How does chain elongation occurs
The OH group on c3 has a nucleophilic attack on the primary phosphate group of another nucleotide. Hydrolysis of the other two phosphates provide energy for the reation
What are the steps to separate the double helix
Helicase opens the helix by breaking the weak h bonds between bases
Topoisomerase releaves the tension in the helix by breaking the dna and reasealing to the right of the replication fork ahead of the helicase
Single stranded dna binding proteins bind to separate strands to prevent annealing
How does dna get a 3’ OH end to start replication
Short rna primer is made by dna primase
How does the antiparallrl nature of dna cause problems
Lagging strand is synthesised discontinuously
What are the discontinuous stands of dna in the lagging strand called
Okazaki fragments
How does replication occurs on lagging strand
Dna primase creates rna primers
Dna polymerase III extends the dna from 3’ OH and makes Okazaki fragments
Dna polymerase I removes RNA primase and fills gaps
Dna Ligase joins discontinuous gaps
What’s in place to prevent dna polymerase detaching
Sliding clamp protein complex
Source of error in dna repl
Frameshifts
Substitution mutations
How are incorrect bases corrected
Dna polymerase has exonuclease activity that can remove the wrong base
What different in the replication of circular dna
2 replication forks as single place of origin
Dna has multipul origins of replication
Base requirements of transcription
Ribonucleoside triphosphates (NTPs)
A template
Rna polymerase
An energy source
How is the correct place in the genome found for transcription
Promoter region containing specific sequences for rna polymerase and regulatory proteins
How does rna polymerase bind to promoter
Sigma subunit between -35 and -10
How does transcription stop
Rna polymerase has a hairpin structure of that are complementary to termination area causing release
What the protein coding region also called
Open reading frame
Transcription in prokaryotes
Proteins coded for that have similar function are located next to each other and transcribed together in a single mRNA called an operon
Features of translation
Code must translate to one of twenty amino acids
Involves ribosomes and tRNA
mRNA read as triplet codons
What are the three stop codons
UAA UAG UGA
How are amino acids attached to tRNA
Aminoacyl-tRNA enzymes
What are the three binding sites in ribosomes
Peptidyl
Exit
Aminoacyl
How is translation initiated in prokaryotes
In the 5’ untranslated region, the shine-dalgarno pairs with the 16S rRNA which places the AUG in the p site of ribosome
A modified methionine is used for the initiating amino acid
How is translation initiated by eukaryotes
The small ribosomal subunit and initiating tRNAi-Met bonds to 5’ cap and scans until AUG
Translation elongation
A charged aminoacyl tRNA enters the A site and pairs with anticodon
Peptide bond is formed between the two amino acids which breaks the covalent bond between the amino acid and tRNA at p-site
The tRNA that was in the p site leaves by e site and the tRNA with the peptide chain moved to P site freeing A site
This repeatedly occurs untill a stop codon enters the a site
Release factors stimulate hydrolysis of polypeptide from tRNA
How is DNA amplified
Polymerase chain reaction (PCR)
Created by Kary Mullis
What process is PCR based on
Dna replication
How many primer are needed for PCR
2 one for each stand of dna
What is the process of PCR
Heated to 96 to denature and break hydrogen bonds
Cool to approximately 55 to allow rna primers and extremophile dna polymerase (taq polymerase) to bond
Heat to 72 for extention
What type of DNA sequencing is used for dna tech
Sanger sequencing
What can be inserted to prevent further extention
Dideoneucleotides because it has only hydrogen on c3 so sugar phosphate backbone can’t form
How does Sanger sequencing work
A mixture of dNTP and ddNTPs are added the ddNTPs with Florescent tags
Dna polymerase extends strand complementary to template
Generated multipul fragments of different lengths different by one nucleotide so the sequence can be discovered
Fragments are separated by size by capillary electrophoresis
