What are nucleic acids?
- Genetic material
- DNA and RNA
What method is used to visualize primary and secondary structures of DNA?
X-ray crystallography
What is DNA? What is it used for?
- Genetic material
- Polymers of deoxynucleotides
- Phosphodiester bonds link nucleotides
- Involved in:
- Info storage
- Gene expression
- Regulation of genetic info
What are the uses of nucleotides in metabolism?
- Drive endergonic reactions (ATP)
- Cell signaling
- Activators (GPCR)
- Secondary messengers (cAMP and cGMP)
- Allosteric regulation of enzymes (ATP, ADP, AMP)
- Phosphorylation of enzymes for regulation
- Functional components of coenzymes
- NAD+/NADH
- NADP+/NADPH
- FAD/FADH2
- Coenzyme A (CoA, CoSH)
What is the primary structure of DNA?
- 1 strand of DNA
- Found in all biological molecules
- Unique order of deoxyribonucleotides (gives protein sequences)
What is the secondary structure of DNA?
- Both strands of DNA
- 2 complementary strands bind together through complementary base pairing
- Antiparallel
What is the tertiary structure of DNA?
Supercoiled/superhelical
What are the 3 types of crystal structures of DNA?
A-DNA (24Å)
- Short and wide
- Right handed
- Dehydrated (doesn’t bind water easily)
- 11 bases/turn
B-DNA (34Å): DNA normally in this form
- Common in cells
- Most stable
- Right handed
- Hydrated
- 10.5 bases/turn
Z-DNA (18Å)
- Saline
- Longest and narrowest
- Left handed
- Next to B form: indicates transcription of the B form will occur
- B and Z can be in the same molecules - sometimes
- Zig zag
- Transient, up to 100 bp
What is DNA denaturation vs renaturation?
Denaturation → single stranded
- “Melting”
- Separation into 2 individual strands
- Replication and PCR
- Transcription
- Causes a hyperchromic shift
Renaturation
- “Annealing”
- 2 DNA strands reform a helix
What is the coding strand? What is the template strand?
Explain base stacking.
- Between adjacent bases in the same strand
- Bases oriented in a plane
- Bases parallel to each other in the same strand of DNA
- Within interior of the helix
- Stability to DNA
- Hydrophobic effect
- Van der Waals
- G-C: more H bonds, more energetically favorable than A-T
What is PCR? What are its steps?
Way to copy specific DNA sequence/gene
1. Double stranded
2. Heated/melted = single strand
3. Cooled to add primers
4. Increase temperature for thermophilic DNA pol and DNA synthesis after primer extension
5. Cool to anneal DNA strands
What is DNA supercoiling?
- Supercoil keeps DNA inside cells
- Structure where majority of DNA molecules inside cell fold upon themselves
- Area where double helix crosses itself
- In prokaryotes
Explain DNA topology.
- Linking number (Lk) = number of times a strand of DNA winds in the right-handed direction around the helix in an imaginary plane
- Twist (Tw) = measures the winding of DNA strands around each other
- Number of turns of B-DNA
- Writhe (Wr) = measures crossing of the duplex of dNA strands
- Lk = Tw + Wr
Compare and contrast DNA and RNA.
- Both store genetic material
- DNA
- Genetic material of most organisms and some viruses
- Inheritance
- Gene expression (template for transcription of RNA)
- Sequences surrounding a gene regulate gene expression
- More stable than RNA since its sugar is missing the 2’OH
- Main genetic material
- RNA
- Genetic material of some viruses
- Gene expression (translation)
- Determines which proteins are present
- Life began with it
- Can act as a ribozyme
- Unstable due to 2’OH
What is a genome?
All the genes that encode proteins
Number of genes increases the more complex the organism is
What is a transcriptome?
RNA transcripts
What is a proteome?
All the proteins expressed by a genome
How is DNA condensed to fit within a cell?
How are chromosomes and telomeres formed?
What are polymorphisms and what are their applications?
- Changes in DNA that could lead to changes in proteins
- Types of polymorphisms
- Single nucleotide polymorphism (SNP): 1 mutation
- Short tandem repeats (STR): deletions and insertions in the genome
- Variable number tandem repeats (VNTR): repeats of a core sequence
- Applications
- Forensic applications: compare DNA sample from crime scene with suspect DNA samples
- Paternity determination
- Precision medicine
What is plasmid-based gene cloning?
Uses plasmids, restriction enzymes, and ligases to isolate genes and express the protein product
What is recombinant technology?
Methods to introduce DNA into bacteria
- Conjugation = transfer of DNA between 2 bacterial cells via direct contact
- Transformation = introduce DNA into bacteria from outside of its cell
- Transduction = viral transfer of DNA to bacteria
What is electrophoresis?
Separation of nucleic acids by size
- Smaller molecules go farther than bigger molecules in the same amount of time
- Ex. SDS-PAGE
