L1: Nucleic Acids & Chemical Bonds Flashcards
what are the two broad areas of molecular biology
- Fundamental principles of the structure and function of DNA, RNA and protein
- Application of molecular techniques to interrogate problems in diverse fields
Gregor Mendel
described genes as “hereditary determinants” but did not know what DNA was
how did scientists answer “what are genes made of?”
- 2 key experiments
1. F. Griffith (1928): pneumonia-causing bacteria
2. Hershey and Chase (1952): bacteriophage T2
what are genes made of? - F. Griffith
- looked at Streptococcus pneumonia
- that bacteria had two strains: rough colonies (non-pathogenic) and smooth colonies (pathogenic)
what are genes made of? - F. Griffith experiments
- injected a mouse with the rough strain
- injected a mouth with the smooth strain
- heat-killed the smooth strain and injected it into a mouse
- mixed the rough strain and the heat-killed smooth strain into a mouse
what are genes made of? - F. Griffith results
- rough strain: mouse was fine
- smooth strain: mouse died
- heat-killed smooth strain: mouse was fine
- mix of heat-killed smooth strain and rough strain: mouse died
what are genes made of? - F. Griffith explanation for results
- in the experiment with the rough and heat-killed smooth strain, there is something at the protein level that made the rough strain turn pathogenic
- Griffith termed the cellular component responsible as “Transforming Principle”
- Griffith believed to be genetic information (found out to be DNA)
what are genes made of?: F. Griffith - why did the rough strain become pathogenic
- the smooth strain has a capsule protecting it
- once heat-killed, the capsule gene is fragmented
- the rough strain then takes in the gene and gets the capsule to protect it
what are genes made of? - Hershey-Chase experiment
- they determined whether genes were comprised of DNA or protein
- used bacterium E. coli and the infecting virus T2 (only has DNA and protein)
what are genes made of?: Hershey-Chase experiment - how the experiment was conducted
- label viruses
- infect bacteria
- agitate cultures
- centrifuge results
what are genes made of?: Hershey-Chase experiment - label viruses
- they grew the virus in the presence of one of two types of radioactive isotopes
- 32P labels DNA
- 35S labels protein
what are genes made of?: Hershey-Chase experiment - agitate cultures
- they knew that the material that the virus injected into the cell was necessary for virus replication
- used agitation to get rid of anything that was not injected into the cells
what are genes made of?: Hershey-Chase experiment - centrifuge solutions
- viral capsids (protein) in solution
- genes/DNA in pellet
what are genes made of?: Hershey-Chase experiment - what does the centrifuge solution imply
- the viral DNA is important since its inside the cells
- the protein is not genetic material since it is not in the cell
structure of DNA - explain what was known about the structure of DNA in the 1950s
- primary structure of DNA is known
- secondary structure is unknown
structure of DNA - what is “Photograph 51”
- it is an x-ray diffraction pattern taken by Rosalind Franklin
- it showed that DNA structure was helical and composed of two strands
structure of DNA - Watson and Crick
- established that
1. DNA is a double helix
2. DNA strands are antiparallel
3. purines (A and G, have 2 rings) pair with pyrimidines (C and U/T, have 1 ring)
structure of DNA : Watson and Crick - what does it mean to be antiparallel
- hydrophilic sugar-phosphate backbone faces exterior
- nitrogenous bases face interior as pairs
- stabilized by hydrogen bonding and hydrophobic interactions
structure of DNA: Watson and Crick - what did the secondary structure show
- a mechanism of DNA replication
- first strand dictates the precise
nucleotide sequence of the other (semiconservative replication)
structure of DNA - what is Chargaff’s rules
- number of purines = number of pyrimidines
- total A = total T, total C = total G
DNA replication - A. Kornberg
- identified DNA polymerase I
- it only works in the presence of a template
- it only synthesizes in the 5’-to-3’ direction
mode of replication - three hypotheses
- semiconservative replication
- conservative replication
- dispersive replication
mode of replication: three hypotheses - semiconservative replication
- parental strands separate and each used as template for the synthesis of a new strand
- result is double stranded molecule with one old (parental) and one new DNA strand
mode of replication: three hypotheses - conservative replication
- parental strands serve as template for an entirely new double helix
- result is original double helix and an entirely newly synthesized double helix
mode of replication: three hypotheses - dispersive replication
- template double helix is broken down into small pieces, DNA replicates, then pieces are brought back together again
- result is two DNA double helices each with new and old segments mixed together
mode of replication - Meselson-Stahl experiment
- determined which hypothesis was correct
- they grew E. coli in the presence of “heavy” nitrogen (15N) to label bacterial DNA
- after many generations (bacterial cell divisions), they moved bacteria onto media with normal nitrogen (14N)
- isolated DNA from the cells after 1 or 2 new cell divisions and separated it based on density
mode of replication: Meselson-Stahl experiment - prediction for gen 0
only 15N band for all hypotheses
mode of replication: Meselson-Stahl experiment - prediction for gen 1
- semiconservative: two hybrid bands
- conservative: one 15N band and one 14N band
- dispersive: two hybrid bands
mode of replication: Meselson-Stahl experiment - prediction for gen 2
- semiconservative: one hybrid, one 14N
- conservative: one 15N, one 14N
- dispersive: both hybrid
mode of replication: Meselson-Stahl experiment - results for all gens
- gen 0: only 15N
- gen 1: hybrids (can rule out conservative)
- gen 2: hybrid and 14N (confirms semiconservative)
mode of replication: Meselson-Stahl experiment - prediction for gen 3
14N band becomes brighter and hybrid band becomes diluted out
DNA vs RNA
- DNA: (1) has a thymine, (2) double stranded, (3)no hydroxyl group at 2’ position but has one at 3’ position
- RNA: (1) has a uracil, (2) single stranded, (3) hydroxyl group at 2’ position and 3’
Francis Crick
- determined DNA secondary structure along with James Watson
- proposed central dogma of biology: DNA -(transcription)-> DNA -(translation)-> protein
- developed adaptor hypothesis
Francis Crick - adaptor hypothesis
- prediction that an intermediate molecule was needed between mRNA and protein synthesis
- bc chemical groups of DNA bases should interact w hydrophilic surfaces but many amino acids have hydrophobic side chains
- adaptor = tRNA
Translation
- read information from mRNA using tRNA to bring codons together to form a polypeptide
- ribosome is needed to catalyze this reaction
define chemical bond
attractive force that holds atoms together
example of chemical bonds
- covalent bonds
- hydrogen, hydrophobic, ionic bonds
chemical bonds - covalent bonds
- strong bond
- do not come apart at physiological temperatures
- electrons are shared between atoms
chemical bonds - hydrogen, hydrophobic, ionic bonds
- weak bonds
- exist transiently, but can be stabilized in large numbers
- based on attractions between electrical charges
weak chemical bonds - hydrogen bond
- forms between positively charged H atom and a negatively charged atom (usually O or N)
- between nitrogen bases so DNA can be pulled apart for replication
- meaningful if you create a ton of them
- Electrons not always shared equally in covalent bond - not full charges
weak chemical bonds: hydrogen bond - how are electrons not shared equally
- atoms with high electronegativity:
1. holds electrons more tightly and is partially negative (δ–)
2. the other atom will become partially positive (δ+)
weak chemical bonds - electronegativity
- O > N > C ~/= H
- oxygen is the most electronegative, followed by nitrogen, then carbon which is approximately equal to hydrogen
weak chemical bonds: electronegativity - polar covalent
electrons are asymmetrically shared
weak chemical bonds: electronegativity - non-polar covalent
- electrons are shared equally
- no partial charges
weak chemical bonds - hydrophobic interactions
- not a true “bond”
- nonpolar groups aggregate to prevent contact with water
weak chemical bonds: hydrophobic bonds - nonpolar vs polar molecules
- nonpolar molecules have no charge and are hydrophobic
- polar molecules have full or partial charges and are hydrophilic
weak chemical bonds - ionic bonds
electrostatic attraction between two oppositely charged groups (full charge)
chemical bonds - high energy bonds
- created through polymerization (a monomer turning into a polymer)
- it’s facilitated by an enzyme and energy is released
chemical bonds: high energy bonds - facilitated by an enzyme
speed chemical reaction rates by lowering activation energies of molecular rearrangements
chemical bonds: high energy bonds - energy is released
- monomers are converted into high energy “activated” precursors prior to polymerization
- these precursors exhibit high energy bonds that yield energy when cleaved
