Mod 3 Flashcards
Pyrimidines vs purines
Pyrimidines
Thymine and cytosine
Single ring
Purines
Adenine and guanine
Double rings
Pure as gold
DNA double helix structure
- 10 base pairs / turn
- Pi-pi interactions as aromatic rings stack next to each other and share electron probabilities
- Major (bp specific) and minor (bp nonspecific) grooves formed
What are nucleic acids composed of?
What does the phosphate do in DNA?
Strong acid
pKa around 1
Makes DNA and RNA acidic molecules negatively charged
Cytosine
- Pyrimidine
- Heterocyclic aromatic ring with 2 substituents attached
1. Amine group (-NH2) at 4’
2. Keto group (=O) at 2’ - Forms 3 hydrogen bonds with guanine
Adenine
- Purine
- Heterocyclic aromatic ring with pyrimidine ring fused to imidazole ring
- Forms 2 hydrogen bonds with thymine or uracil
Uracil
- Only in RNA
- Pyrimidine
- Heterocyclic aromatic ring
- Forms 2 hydrogen bonds with adenine
Thymine
- Only in DNA
- Pyrimidine
- Heterocyclic aromatic ring
- Forms 2 hydrogen bonds with adenine
Guanine
- Purine
- Heterocyclic aromatic ring with pyrimidine ring fused to imidazole ring
- Forms 3 hydrogen bonds with cytosine
What process allows for the degradation of the covalent backbone of RNA and DNA through hydrolysis?
Hydrolysis
- Slow, non-enzymatic hydrolysis of phosphodiester bonds.
Faster in RNA due to the 2’ OH group, which makes RNA more reactive than DNA.
Cyclic 2’,3’ monophosphate nucleotides are produced first and are further hydrolyzed into 2’- and 3’- nucleoside monophosphates.
How is condensation related to DNA synthesis?
- 3’ OH of growing chain + 5’ OH of deoxynucleoside monophosphate
- Thermodynamically unfavorable
- Coupled with favorable reaction to proceed (using dNTP)
Role of dNTP in DNA synthesis?
- High-energy triphosphate at 5’ end
- Hydrolysis to dNMP + pyrophosphate drives condensation
- Net negative ΔG when coupled
DNA structure?
- Double helix, 2 antiparallel strands
- H-bonds between complementary bases
- Phosphate-deoxyribose backbone (hydrophilic)
- Bases stacked inside, regular major/minor grooves
What did Erwin Chargaff discover?
- Mole percent A = T
- Mole percent C = G
- Due to complementary base pairing
Who discovered DNA structure?
Dr. James Watson & Francis Crick
Difference between nucleoside and nucleotide?
Nucleoside: no phosphate
Nucleotide: with phosphate
Primary structure of protein?
- Linear amino acid sequence
- Peptide bonds link amino group and carboxyl group
- Sequence determines protein shape and properties
Secondary structure of protein?
- Alpha helices or beta sheets
- Stabilized by hydrogen bonding patterns
Tertiary structure of protein?
- Globular shape
- Hydrophobic residues inside, away from water
- Essential for protein function (e.g., hemoglobin holding heme)
Quaternary structure of protein?
2+ polypeptide chains form one molecule
Example: Hemoglobin has 4 subunits working together
Examples of proteins and their functions?
Antibodies: defense, bind foreign molecules
Collagen: structure, triple helix
Calcium pump: transport, ion movement for muscle contraction
Insulin: communication, regulates blood sugar
Alpha amylase: catalysis, breaks down carbs
Ferritin: storage, stores iron
What is chirality in molecules?
- Molecule’s reflection can’t be superimposed on the original
- Central carbon attached to 4 different groups
Difference between L and D chirality?
- L chirality (levo): rotates light left, typical in cells
- D chirality (dextro): rotates light right
Which amino acid is achiral?
Glycine
R group is hydrogen and there is already a hydrogen
What is a peptide bond?
- Covalent bond between carboxyl and amino groups
- Formed through condensation
- Rigid, planar due to resonance
Ends of a polypeptide chain?
Amino (N) terminal: beginning
Carboxyl (C) terminal: end
Nonpolar, aliphatic amino acids?
Met, Pro, Gly, Ala, Val, Leu, Ile
- Hydrophobic
- Cluster in proteins (hydrophobic effect)
Pro: rigid ring, found at protein surface
Polar, uncharged amino acids?
Ser, Thr, Cys, Asn, Gln
- Can form H-bonds
- Often on protein surface (interact with water)
Aromatic R group amino acids?
Phe, Tyr, Trp
- Hydrophobic, absorb UV light at ~280 nm
- Used to measure protein concentration
Phe: most hydrophobic
Tyr: can form H-bonds
Trp: bulky, aromatic
Negatively charged R group amino acids?
Asp, Glu
- Negative charge at pH 7
- Hydrophilic
- Found on protein surface, in contact with water
Positively charged R group amino acids?
Lys, Arg, His
- Basic, polar
- Found on exterior or enzyme active sites
Histidine: least basic (imidazole ring)
Peptide bond formation
- Nucleophilic attack from electron pair on the amino group on one aa to the carbonyl of the other aa (nuc addition)
- Release of a water molecule (elimination)
What is the purpose of SDS gel electrophoresis?
Resolves proteins according to their molecular weight.
How does SDS denature proteins in SDS gel electrophoresis?
- Anionic SDS detergent is added to denature the sample.
- SDS binds to proteins, causing unfolding
What effect does SDS have on proteins during electrophoresis?
- SDS molecules uniformly coat proteins with a negative charge.
- Results in similar shape and mass-to-charge ratio.
How does SDS gel electrophoresis separate protein subunits?
SDS disassociates protein subunits into individual polypeptide chains.
What happens after the protein sample is prepared in SDS gel electrophoresis?
- Mix transferred into a well at the top of a polyacrylamide gel.
- Negatively charged polypeptides move toward the positive end.
Why do smaller polypeptides move faster in SDS gel electrophoresis?
- Smaller proteins face less resistance in the gel.
- Speed depends on molecular weight, as mass-to-charge ratio is similar for all.
What is the first and last amino acid found in the body?
Asparagine
Threonine
How are amino acids classified?
- 20 common alpha amino acids
- Chiral center
- Optically active (L and D enantiomers)
- Differ based on R groups (side chains)
Who devised the one-letter code for amino acids?
- Margaret Oakley Dayhoff
- For space efficiency in bioinformatics
What is the significance of the letters CHIMSV in the amino acid code?
First letter unique for each amino acid.
What is the Greek letter naming convention in amino acids?
- Alpha carbon is the center
- Beta (β), Gamma (γ), Delta (∆), Epsilon (ε)
What is the number naming convention in amino acids?
C1 is the carbon with the highest atomic number substituent
Usually carbon in COO-
Then numbers continue to the other end
What is the absolute configuration of amino acids based on?
- L-glyceraldehyde
- L = levorotatory (left)
- L and D refer to absolute configuration, not optical properties
Why are most amino acid residues L?
Bc active sites of enzymes are asymmetric and therefore reactions are stereospecific
Rare D-configuration ones are introduced via enzyme-catalysed reactions that occur after ribosome synthesis
How are amino acids grouped based on polarity? (5)
- Nonpolar, aliphatic
- Aromatic
- Positively charged
- Negatively charged
- Polar, uncharged
What does the Lambert-Beer Law relate to?
- Absorption of light by biomolecules at characteristic wavelengths
- Determines concentration using a spectrophotometer
What are some examples of other amino acids formed postsynthetically?
4-hydroxyproline (protein derivative in collagen)
γ-carboxyglutamate (in prothrombin, Ca2+ binding)
Desmosine (in elastin, derivative of 4 lys residues)
Selenocysteine (derived from serine)
Pyrrolysine (in methanogenic archaea)
Ornithine, Citrulline (precursors for arginine)
How can protein activity be modified?
Addition of phosphoryl, methyl, acetyl, adenylyl, ADP-ribosyl, etc.
What ionization behavior is seen in amino acids?
3 things
- Ionizable R groups, amino, and carboxyl groups
- Amphoteric: can act as acid or base
- Zwitterion: dipolar ion
What happens during the titration of glycine with NaOH?
First stage: COOH loses H
Midpoint: equal concentrations of proton donor (COOH) and proton acceptor (COO-)
Point of inflection: pKa = pH of protonated group
Second stage: NH3+ group loses proton
What is the isoelectric point (pI) of glycine?
pI is the mean of two pKa values
No net charge when at pI, glycine is fully ionized and dipolar
What determines the titration curve of amino acids with ionizable R groups?
- More complex with 3 stages
- Isoelectric point depends on the R groups present
- Two carboxyl groups lower the pI, two positively charged groups raise the pI
Which aa provides significant buffering power near neutral pH?
Histidine (due to its ionizable R group)
What is a peptide bond?
Amide linkage formed between two amino acids
Dehydration reaction: OH from one carboxyl and H from another amino group
What is the reverse of peptide bond formation?
Hydrolysis (hydrolytic cleavage)
Favored under biological conditions
What is the difference between a dipeptide, tripeptide, and polypeptide?
Dipeptide: 2 amino acids
Tripeptide: 3 amino acids
Polypeptide: long chain of amino acids
What is a protein versus a polypeptide vs an oligopeptide?
Polypeptide: molecular weight < 10,000
Protein: molecular weight > 10,000
Oligopeptide: a short polymer of amino acids joined by peptide bonds
How does the ionization of amino acids change when they become peptide residues?
Chemical environment of the residue alters pKa values
Ionizable R groups may have different pKa values
3 example of small biologically active peptides?
Oxytocin: 9 residues, stimulates uterine contractions
Thyrotropin-releasing factor: 3 residues, stimulates release of thyrotropin
Amanitin
What are multisubunit proteins?
Proteins with 2+ polypeptides, can be identical (oligomeric)
What are protomers
Identical subunits
Hemoglobin: 4 subunits, 2 alpha, 2 beta
dimer of ab protomers
How to estimate # of aas
Can estimate the # of residues by dividing its molecular weight by 110
- Assuming no other chemical constituents
- Considers the higher frequency of smaller residues and the impact of water’s weight
What is a prosthetic group?
- A metal ion or organic compound bound to a protein
- Essential for its activity
- Found in conjugated proteins
What are conjugated proteins?
- Proteins with one or more prosthetic groups
- Classified by the chemical nature of the prosthetic group
