Biochem Exam 2 Flashcards
What is the primary structure of a protein?
Amino Acid sequence and location of S-S bonds
What is the main bonding interactions for primary structures?
Covalent Bonds
What kind of covalent bonds are found in proteins?
Amide bonds and disulfide bonds
What is secondary protein structure?
Interaction of amino acids close to each other to form specific patterns
What is the main bonding interaction for secondary protein structure?
Hydrogen bonds
What is the main bonding interaction for tertiary strucutre?
Hydrophobic effect
What is Quaternary Structure?
Arrangement of multiple folded protein molecules (subunits) in a multi-subunit protein.
What is the main bonding interaction in forming quaternary structure?
Hydrophobic effect
Are there usually disulfide bonds or covalent bonds between subunits?
No
What percent Double Bond are peptide bonds? What does this make them?
40% double bond character makes them planar
Are peptide bonds usually cis or trans?
Trans
Can you rotate around a peptide bond?
No
Who discovered the nature of the peptide bond?
Pauling and Corey
Where can you rotate bonds in the protein chain?
Around the alpha carbon
What are the angles on either side of the alpha carbon called?
Phi an psi angles
What is the angle of the phi and psi bonds in a fully extended chain?
Close to +/- 180°
Why are certain values of phi and psi forbidden?
Certain values are forbidden because of steric hindrance
What is a Ramachandran plot?
Calculated plot of all sterically allowed psi and phi angles for a polypeptide chain
What does the green space on the Ramachandran plot represent?
Angles of phi and psi where Glycine is present
What does the blue space represent on the Ramachandran plot?
The angles of phi and psi for the other 19 amino acids
What is the definition of secondary structure?
The general 3D form of local segments or biopolymers such as proteins and nucleic acids (DNA/RNA) to give regular, recurring local conformations
What are general secondary structures in proteins?
alpha helix, beta strand, beta bend, collagen triple helix, loops & turns
What do the Hydrogen bonds occur between in protein secondary structure?
backbone amide NH of one Amino Acid and backbone carbonyl C=O of another
Who proposed the alpha helix?
Linus Pauling
Which residues does hydrogen bonding occur between?
n and n+4 residues
What residues bind in a 3-10 helix?
n and n+3
Who are considered the founders of structural biology?
Linus Pauling & Robert Corey
Which way do R groups point in alpha helix structures?
R groups point outwards, perpendicular to axis
Describe the orientation of Hydrogen bonds relative to the alpha helix
H-bonds are parallel to helix axis
What is the pitch on an alpha helix?
5.4 angstroms
How many residues does it take to make one turn of the alpha helix?
3.6 residues per turn
What is the rise of the alpha helix?
1.5 angstroms
Are alpha helices in nature right or left handed?
Right handed
Does an alpha helix have a dipole? Why?
Yes, alpha helices have dipoles. This is because all C=O groups point towareds the C-terminus, giving entire helix a dipole with (+) N, (-) C-termini
Do stable alpha helices typically end with a (+) or a (-) charged amino acid? Why?
Stable alpha helices typically end (C-term) with a (+) charged AA to neutralize the dipole moment.
Is proline found in alpha helices? Why?
Proline is not likely found in alpha helices because its cyclic side chain sterically destabilizes an alpha helix
What causes steric repulsion in alpha helices?
Steric (/charge) repulsion by adjacent bulky (/like-charged) amino acids
Which AAs are too large to be found in alpha helices?
Tryptophan and Tyrosine
Which AAs are too small to be found in alpha helices?
Glycine
Can alpha helices be amphipathic? What doe this mean?
Alpha helices can be amphipathic - This means that hydrophobic resides on one side and hydrophilic residues on opposite side
What is alpha-Keratin?
A coiled-coil motif
What is alpha-keratin made up of?
Amphipathic alpha helices
Where is alpha-Keratin commonly found?
Hair, skin, and nails
What is alpha Keratin rich in? Why is this signficant?
Alpha-Keratin is rich in Cysteine, which is important because Cys forms -S-S- bonds
What is the biochemistry behind getting a perm?
Heat reduces -S-S- bonds, which breaks linkages; Next, curl; Then re-oxidize to form -S-S- bonds; This holds hair in new position
What can alpha keratin be used to detect? Why does this work?
Alpha-keratin can be used to detect heavy metal poisoning. This is because Cys attracts strong metals.
What are Beta Stands
Secondary structure in which polypeptide strands are almost fully extended
How many B strands are in B sheets?
At least two Beta strands are required to make a Beta sheet
Where can strands in Beta sheets come from?
Stands can come from same polypeptide, completely different sections of a polypeptide, or even different polypeptides.
What plane are strands in in Beta sheets?
Strands are in X-plane
Where are H-bonds oriented in Beta sheets?
H-bonds occur between amino acids on adjacent strands, perpendicular (Y direction) to strand direction
Where are R groups oriented in Beta strands?
Side chains (R-groups) alternate in pointing above and below plane, perpendicular to H-bonds AND strands (Z plane)
Why can Beta sheets accomodate more bulky substituents?
Because the R groups alternate in pointing above and below the plane of the sheets. This limits adjacent bulky/like charged residues.
What is the difference between parallel and antiparallel Beta sheets?
In Antiparallel sheets, strands run in opposite N- to C-terminal direction, while in Parallel sheets, strands run in the same N- to C- direction
Is Anti-parallel or parallel Beta sheets stronger?
Antiparallel are stronger because they give you linear H-bonds
What is the general overall structure of a Beta strand?
Extended zig-zag (ruffled) conformation
How many residues are involved per strand in Beta sheets?
6-12 residues involved per strand
What is the order of AAs pattern in Beta Sheets?
Repeating units of 2 AA residues with distance = 7 angstroms (3.5 A per AA)
Can Beta sheets be amphipathic?
Yes; One surface of Beta sheet may consist of hydrophobic side chains (R grps) while other can have hydrophilic ones
Do Beta strands in a Beta sheet twist?
Yes, Beta strands can twist
Can some proteins be all Beta sheets?
Yes
What is an example of Beta Sheets in nature?
Spider’s silk
What makes spider silk so strong?
Extended layers of antiparallel Beta Sheets
What is the AA pattern in spider silk?
(-Gly-Ser-Gly-Ala-Gly-Ala-) - Alanine from 1 sheet interdigititates with Alanine from another sheet.
What is collagen notably used for?
Collagen is the main structural protein in the extracellular matrix (ECM) in connective tissue in animals
What percentage of protein mass in large animals is made up of collagen?
Roughly 1/3rd of the total protein mass
What is collagen present in?
Bone matrix, tendons, cartilage, blood vessels, skin, etc.
What form is collagen?
Triple helix
Which AA is found abundantly in the collagen triple helix?
Proline
What is the general repeating AA pattern of the collagen triple helix?
G-P-P/Hyp
What residue is invariant in the collagen triple helix?
Glycine - It is located along central axis of triple helix (other residues cannot fit)
Where is H-bonding located in collagen triple helix? What residues does it bind between?
H-bonding is interchain and occurs between amide NH of one helix to carbonyl oxygens of another helix
Are there any intrachain H-bonds in collagen triple helix?
No
What is the cause of one type of Osteogenesis Imperfecta?
Occurs when a mutation in a gene for collagen leads to the substitution of Glycine for another amino acid. This prevents the normal production of mature collagen, which leads to brittle bones.
What structure is this?
Ascorbic Acid (Vitamin C)
How does Vitamin C help with building the collagen triple helix?
Formation of HydroxyPro and HydroxyLys requires oxygen and vitamin C
What is a vitamin C deficiency called?
Scurvy
What are the effects of scurvy?
Scurvy results in a lack of hydroxyPro, which leads to weak collagen (skin lesions, fragile blood vessels, bleeding gums)
What are collagen fibrils strengthened by?
Intrachain lysine-lysine and interchain hydroxypyridinium covalent crosslinks
What are the effects of collagen cross-links as they accumulate over time?
Increase in cross-links lead to less elastic collagen, which leads to more brittle bones/tendons (many of the signs of old age)
What kind of secondary structures are loops and turns?
Non-regular secondary structures
What do turns allow?
Turns and loops allow chain reversal
Where are loops are turns found?
Normally on surface of globular proteins (hydrophilic)
What does the Beta turn allow?
Allows polypeptide in antiparallel Beta sheet to reverse direction
What residues are bonded in a Beta turn?
Carbonyl O’s are H-bonded to amide H’s between residues 1 and 4
Which amino acid helps make the tight, rigid turn as the 2nd residue in the turn?
Proline
Where are Beta sheets found in the Ramachandran plot? Why?
Beta sheets are found in the top left corner of the Ramachandran plot, because Beta sheets have elongated strands, so phi and psi angles are close to 180°
What is tertiary structure? What causes tertiary structure to form?
Tertiary structure results from the folding of a polypeptide chain into a closely-packed 3D structure, mainly because of hydrophobic effect
What is one effect of tertiary structure?
Amino acids far apart in the primary structure may be brought closer together
What does X-ray crystallography do?
Reveals 3D structure in atomic detail
What do you need to perform x-ray crystallography?
Crystals of the protein (difficult to obtain)
What is one caveat with x-ray crystallography?
It gives a frozen picture of proteins (when they are “alive”, proteins move)
What are the steps to calculating protein structure from the diffraction pattern of x-ray crystallography?
- Obtain diffraction pattern
- Fourier Transform
- Computer fit map to protein sequence
- Fixed (frozen) picture of protein
Why is X-ray crystallography difficult to perform for many proteins?
You have to have protein crystals, which are very difficult to get for many proteins
What new technique can be used to visualize proteins in solution?
2D NOESY NMR Spectroscopy
What are 3 limitations of the NOESY technique for visualizing proteins?
- Need high concentrations of protein
- Time consuming
- Only works for small proteins
What does it mean for proteins to be “dynamic” in solution?
Their structure can be displaced up to 2 angstroms
What is Cryo-Electron Microscopy? (CEM)
Form of transmission electron microscopy (TEM) where the sample is studied at cryogenic temperatures
What do you do with a protein structure once you have determined it?
Add it to a protein data bank
What is the difference between globular proteins and fibrous proteins?
Globular proteins are water soluble, while fibrous proteins are normally insoluble
What causes proteins to denature?
Heat
Can all proteins be refolded?
No
What happens to proteins that unfold within a cell? What molecule carries this action out?
They are tagged for destruction by Ubiquitin
What is the name of the complex that “shreds” proteins?
26s Proteosome
What is the term for protein degredation?
Proteolysis
What are chaotropic reagents? What do they do? How do they work?
Proteins are denatured by Chaotropic agents; They work by disrupting interactions that stabilize tertiary structure
Who is Christian Anfinsen? What is he known for? Describe his experiment.
Anfinsen is known for discovering that primary structure determines tertiary structure. He did this by using 8 M Urea and mercaptoethanol to denature RNase A (a protein that does refold, luckily). Whenever he dialyzed out Urea and Mercaptoethanol, the protein refolded, because the primary structure was still in tact.
What groundbreaking discovery did Christian Anfinsen make?
He showed the primary structure determines tertiary structure
What determines the tertiary structure in proteins?
Primary structure
What are the interactions that help maintain tertiary structure? (5)
Hydrophobic effect, ionic interaction, metal ion coordination, hydrogen bonds, and disulfide bridges
What factors of protein folding are (-) deltaG (favorable)? What about (+) deltaG (unfavorable)?
The internal reactions involving deltaH (enthalpy) are favorable, along with the deltaS (entropy) of the water molecules in the hydrophobic effect ….. deltaS (entropy) of the folded protein is unfavorable.
What must be overcome for proteins to fold? (energetics)
The large decrease in entropy (deltaS) of the folding protein
How long does it take for most proteins to fold?
Less than 10 seconds
What does Levinthal’s Paradox state?
Protein DON’T sample all possible folding conformation & must only sample through limited conformations, & thus fold be “specific pathways”
What is the order of steps in which proteins fold in the accepted model of protein folding?
- The protein collapses in upon itself due to hydrophobic effect
- Secondary structures quickly form
- Then, the tertiary structure begins to form
- Finally, the native state forms