Biochemistry 1 Flashcards
Amino Acid Backbone
Alpha carbon, amine group, carboxyl group, side chain (R group).
Amino acids are amphoteric.
At low pH both the amine and carboxylic acid are protonated.
- NH3+ pKa 9.5
- COOH pKa 2
Above pH of 2, but below 9.5: COO- and NH3+ (zwitterion).
Above pH of 9.5: COO- and NH2
AA Acidity and Basicity
AA are characterized as acidic or basic based on the pKa of their side chain (R group)
-Acidic if R group contains a carboxyl group; basic if R group contains amine
When the pH of solution=pKa of the side chain, half of the AA that R group protonated, and half have it deprotonated; as pH rises, the amount of AA with the R group deprotonated increases towards 100%.
pKa’s to know
- Asp (4)
- Glu (4)
- His (6.5)
- Lys (10)
- Arg (12)
Hydrophilic or Hydrophobic
Amino acids are characterized as hydrophilic (polar) or hydrophobic (non-polar) based on their side chains.
Non-polar/hydrophobic: R group is alkyl or aromatic.
Polar/hydrophobic: acidic R groups, basic R groups, and other R groups that contain +1 very polar bond(s) in R group.
Isolelectric Point
pH at which molecule is net neutral.
-for basic AA: PI=
(pKa R group + pKa amine)/2
-for acidic AA: PI=
(pKa R group + pKa COOH)/2
-for neither acidic nor basic AA PI:
(pKa COOH + pKa amine)/2
-> (9.5+2)/2 => 5.75
-must have NO pKa of side chain (R group cannot be protonated or deprotnated).
Isoelectric Focusing
Separate proteins based on their isoelectric point (pI)
-pH gradient set up in gel (low pH is + end, high pH is negative end).
- Proteins start at either end and migrate towards the opposite end until they reach the position in the gel’s pH gradient that is equal to their pI.
- Protein starting at low pH end will be net + and repelled by positive charge at that end; migrates towards opposite end (- and high pH) until it reaches pI, at which point if travelled further it would begin to have a net -charge and be repelled.
Peptide bond
Amide linkage between AAs in a polypeptide (protein).
Peptide bond has resonance: increased stability of bond (difficult to hydrolyze).
-6 atoms (those of amide bond and those directly adjacent) are planar due to partial pi bond character.
Formation of the peptide bond
condensation reaction (remove H20; also called dehydration) and is facilitated by tRNA molecules during translation.
- Addition-elimination reactions between carboxylic acid and primary amine (amine attacks the carbonyl carbon).
- Reaction is non-spontaneous (delta G>0) and requires catalysis and ATP.
Breaking peptide bond
a hydrolysis reaction.
-Requires strong base and enzyme catalysis.
Sulfur Linkage
Disulfide bond/bridge: covalent bond between sulfurs of two cysteine residues (AAs called residues when in a polypeptide)
R-SH + R-SH -> R-S-S-R
(oxidation)
cystine
“cystine” =2 linked cysteine
residues.
Forms when cysteines are close by each other and in an oxidizing environment
- Cytosol is a reducing environment, so no disulfide bridges there.
- They can form in RER lumen, secreted proteins, proteins on cell membrane exterior.
Protein structure: Primary structure
The sequence of covalently linked AA in a polypeptide chain.
-Held together by peptide bonds.
- Only broken by hydrolysis reaction during catalysis.
- Proteases degrade peptide bonds.
- Determined by DNA sequence of gene
Protein structure: Secondary structure
Local regions of folding of the polypeptide chain due to interactions between backbone atoms.
-Hydrogen-bonding between backbone atoms: H atom of amino group is the H-bond donor and O of carbonyl group is the H-bond acceptor.
- Most common secondary structures: alpha helix and beta pleated sheet.
- Protein may contain one, both, or neither motif.
- Amino acids have different propensities for forming alpha helices and beta sheets.
Alpha Helix
Helical structure formed by H-bonds that run parallel to the axis of the helix and form between every 3-4 AAs.
Right-handed helix with approximately 3.6 residues per turn of the helix.
Forms within one continuous region of a polypeptide chain.
R groups stick outwards from helix (not inwards; not enough space).
Alpha helix “breakers”: proline (cyclic R group sterically hinders helical shape) and glycine (moves very freely because R group is so small).
Beta pleated sheet
2 or more different segments of a polypeptide chain align and H-bonds between adjacent strands form perpendicular to the length of the chain.
The aligned sheets are pleated at the alpha C of the backbone.
R groups just out above and below the sheet.
Large aromatic residues and large alkyl residues favored (tyrosine, tryptophan, phenylalanine, isoleucine, etc.)
Parallel sheet
N-terminus of one sheet
aligns with N-terminus of the other.
Anti-parallel sheet
N-terminus of one aligns with C-terminus of the other (strands run in opposite directions).
Tertiary Structure
The overall 3D shape that a polypeptide chain folds into due to interactions between side chains.
Involves mainly non-convalent bonds:
-London dispersion forces between non-polar side chains.
- Dipole-dipole between polar side chains.
- H-bonds between polar side chains with H-bond donors and acceptors.
- Ionic bonds between charged side chains (acidic/basic): salt bridges.
Disulfide bonds also considered part of tertiary structure.
Tertiary structure is the conformation that is most stable/lowest energy by maximizing IMF.
Polar aqueous environment
Physiologic Conditions.
Hydrophobic AAs cluster together on the interior region of the protein and polar residues are exposed on the exterior of the protein.
-Enables polar residues to have H-bonding or dipole-dipole interactions with the surrounding water molecules.
Hydrophobic Interactions
London dispersion forces: are weak individually but present in such a high quantity that they contribute immensely to the overall structure and stability of a folded protein.
- This is the main force driving the folding of a protein.
- Favorable because it maximizes the entropy of the folded protein: enables water molecules to engage in a maximum amount of highly dynamic and disordered H-bonds (rather than being trapped in a highly ordered solvation layer in which water molecules interact weakly with exposed non-polar side chains.
Quaternary Structure
Multiple subunits (folded polypeptide chains) interacting.
- Only found in proteins that are composed of more than one polypeptide chain.
- Ex. Hemoglobin has 2 alpha subunits and 2 beta subunits.
- Held together by same interactions as tertiary structure (disulfide bridges, London dispersion, dipole-dipole, H-bonds, ionic bonds/salt bridges).
Protein Folding
The protein’s primary structure and the environment it is in (polar versus non-polar) determine the shape the protein will fold into.
- Under physiologic conditions a protein will fold into its native conformation, which is usually biologically functional.
- Lowest energy state
- Protein may have more than one native state possible.
-Proteins begin folding as they are translated by ribosomes.
Denaturation
Process by which a protein is unfolded from its native state.
- Secondary, tertiary, and quarternary structures are lost (primary isn’t).
- Denatured protein can no longer perform its proper function.
- Denaturation can occur via changes in: temperature, pH (adding an acid or base), or chemical environment (adding a salt or dramatically changing the polarity of the environment by adding some organic solvent).
- Increases in temperature will generally increase protein activity until the temperature of denaturation is reached, at which point it becomes non-functional.
- Usually denaturation is reversible.
- Primary sequence directs protein to refold into the same conformation.
- Chaperones may or may not be required.
Temperature
Molecules move around so rapidly that they break free from ordered structure
pH
Salt bridges and H-bonding disrupting by changing residues’ protonation state
salt
Disrupts ionic bonds, H-bonds, dipole-dipole
Solvent
Disrupts H-bonds and dipole-dipole
Enzymatic Proteins
Enzymes: protein catalysts -> increase the rate of chemical reactions by lowering the reaction’s activation energy.
Lowers the activation energy by stabilizing the transition state.
Enzymes may participate in the reaction through transient bonds or IMF, but comes out unchanged.
Substrate: compound the enzyme acts
on -> enzyme has a specific substrate and only catalyzes a specific reaction (or a few closely related reactions) for that substrate.
Enzyme names usually end in “-ase”, and the prefix may indicate what substrate it is. (ex. protease, amylase, nuclease)
Classes of enzymes
Oxidoreductase Transferase Lyase Hydrolase Isomerase Ligase/Synthetase
Oxidoreductase
catalyze redox reactions (transfers of electrons). Ex: Dehydrogenase.
Transferase
catalyze reactions in which a group of atoms is transferred from one substrate to another. Ex: Kinase.
Lyase
Catalyze reactions in which functional group is added, breaking a double bond (or the reverse). Ex: Aldolase.
Hydrolase
catalyze hydrolysis reactions (break a molecule with addition of water or form a molecule with removal of water). Ex: Lipase.
Isomerase
move atoms around on one molecule so that it changes into a new isomer. Ex: phosphohexoisomerase.
Ligase/Synthetase
catalyze reactions in which two substrates are joined new (new C-C,
C-O, C-S, or C-N bond) and reaction coupled to ATP hydrolysis. Ex: DNA ligase
Binding proteins
proteins have a specific affinity to the molecule that they act on.
- Affinity: how strongly the protein is attracted to/interacts with its target molecule.
- Proteins on the surface of the cell membrane bind to specific substances in the external environment to relay a signal to the inside of the cell or to interact with other cells in the environment.
Non-enzymatic proteins
Have many functions, including transportation, regulation, structural, hormonal, and defense.
Immune system proteins
Antibodies are proteins in the immune system that detect foreign substances (antigens) and mark them for destruction.
-Antigen-antibody specificity is crucial.
Motor proteins
Enable movement of the entire cell or certain substances with the cell.
- Myosin in muscles enables them to contract.
- Kinesin and dynein use ATP to move along microtubule filament “tracks” and transport subtance; dynein also involved in cilia and flagella movement.
