Biochemistry Quiz 2 (lectures 5-8)

Protein Structures are stabilized by _____

Noncovalent interactions and forces

Tertiary Structure

Well-defined 3-D fold of a polypeptide

• weak interactions and covalent bonds hold interacting segments in position

• determined by the amino acid sequence of the protein

Primary Structure

covalent bonds linking amino acid residues in a polypeptide chain

Secondary structure

recurring structural patterns-describes the spatial arrangement of the main-chain atoms in a segment of polypeptide chain

quaternary structure

2+ polypeptide subunits/chains in 3D complex

What do ribosomes use to join amino acids together into proteins? (backbone)

mRNA code

The peptide bond is _____ and ______ (shape)

rigid and planar

phi φ (dihedral) angle bonds:

N and Ca

psi ψ (dihedral) angle bonds:

Ca and C

omega ω (dihedral) angle bonds:

C and N

What resonates in the peptide bond

carbonyl oxygen and amide nitrogen

The partial negative and positive charge in a peptide bond set up a _______ which leads to _________

small electric dipole; partial double bond character (and little rotation)

Architecture of peptide bonds

• Cannot rotate freely

• 6 atoms of peptide group lie in single plane

• partial double bond character of C-N peptide

• Bond prevents rotation, limiting conformation range

What defines peptide conformations

Dihedral angles

Why is there very little rotation around the peptide bond

The partial double bond character that makes the peptide bond planar

regular secondary structure

φ and ψ remain the same throughout the segment

common types of secondary structure:

alpha helix, beta conformation, beta turn, random coils

what are secondary structures stabilized by

noncovalent interactions: H bonding, hydrophobic interactions, van der wals forces, disulfide bonds

alpha helix

• simplest arrangement of secondary structure with max number of H bonds (common) and backbone wound around an imaginary axis

• tough insoluble protective structures of varying hardness and flexibility

Each helical turn in an alpha helix has:

• 5.4 Å

• 3.6 residues

A hydrogen bond is formed in an alpha helix between ____ and _____

the electronegative Nitrogen of residue n; the electronegative carbonyl Oxygen atom of n+4

What bond provides significant stability for the alpha helix

Hydrogen bonds between main chain atoms: N-H and O=C

What is important to the function of an alpha helix protein and interacting partners (helical wheel)

The position and type of R group

What disrupts an alpha helix

Proline and Glycine

• Proline: introduces a destabilizing kink, N atom in rigid ring, and rotation about N-C alpha is not possible

• Glycine: H atom is the R-group, too much flexibility to stabilize

What handedness is the alpha helix (and why)

Right handed:

• R groups protrude away from helical backbone

• most common

• more stable (than left which isn’t observed in proteins)

• small electric dipoles in peptide bond align through H bonds

Increasing number of residues does what to the helical radius of an alpha helix

increases helical radius (direct relationship): you can find helix type from number of residues

What is a residue

An amino acid in an alpha helix

Beta Conformation

• backbone extends into zigzag

• organizes polypeptide chains into sheets with the R group extending out of the plane of the sheet

• soft and flexible filaments

Beta strand

single protein segment in beta conformation

beta sheet

several strands in beta conformation side by side

Antiparallel beta conformation

• more frequent

• opposite orientation (direction)

• more stable because H bonding is in line (linear)

Parallel beta conformation

• same orientation

• occur less frequently

• less stable because H bonding is at a distorted angle

What bonds form between adjacent segments in Beta Sheets

Hydrogen bonds between backbone atoms

Beta Turns

• connect ends of two adjacent segments of antiparallel beta sheet

• 180-degree turn

• involves 4 residues

• hydrogen bonding forms between the first and fourth residue

Type 1 Beta turn

proline is residue two (causes a kink)

type 2 beta turns

glycine is residue 3 (often occuring)

gamma turns

involve 3 residues: 180-degree turn and proline on residue 2

What defines Protein Conformations

dihedral angles

What feature describes the secondary structure of a protein

dihedral angles phi and psi associated with each residue-Ramachandran plots visualize these

What amino acid is not typically shown on a Ramachandran Plot

Glycine (typically falls outside of expected range)

Circular Dichroism

• Assess secondary structures

• measures differences in the molar absorption of L vs R handed circularly polarized light

Chromophore

Peptide bond

Most common AAs for alpha helix

KHMLACEQ (Kris Has Marvelous LACE Q-tips)

Most common AAs for beta sheet

IVYFTW (IVY For The Win)

Most common AAs for reverse turns

SPDNG (SPeeDiNG)

Major types of protein groups based on polypeptide chains

fibrous, globular, membrane, intrinsically disordered

fibrous proteins

arranged in long strands or sheets

• some can be adapted for structure:

  • give strength

  • simple repeating element of secondary structure

  • H2O insoluble due to high hydrophobic residue concentrations

• 
  

globular proteins

folded into spherical or globular shape

membrane proteins

embedded in hydrophobic lipid membranes

intrinsically disordered proteins

lacking stable tertiary structure

Structure of alpha-Keratin

• R-handed alpha helix

• Two strands of alpha keratin in parallel and wrap around each other to create coil

• super-twisted helical path is left-handed

• Points of contact are rich in hydrophobic residues: A, V, L, M, P

• Cross links are stabilized by disulfide bonds

Structure of Collagen

(found in connective tissue)

• secondary structure-left handed alpha chains with repeating tripeptide unit (often G, P, and 4-Hyp)

• tertiary and quaternary structure: r-handed twisting of 3 separate polypeptides

Structure of Collagen Fibrils

• Cross-linked by covalent bonds (Lys, HyLys, or His)

• links created by uncommon amino acid residues

Vitamin C

required for hydroxylation of Proline and Lysine in collagen-scurvy can degenerate connective tissue

Globular Proteins

• Fold back on each other

• more compact than fibrous proteins

• enzymes, transport proteins, motor proteins, regulatory proteins, immunoglobins

• Each has distinct structure for function (ie Myoglobin)

Intrinsically Disordered Proteins

• lack definable structure

• lack hydrophobic core

• high densities of charged residues (K, R, E,) and Proline

• facilitates a protein to interact with it for multiple binding partners

• intrinsically disordered segments can assume different structures

• anything above 0.5 PONDR score is not folded and anything below is

alpha beta barrel

series of beta-alpha-beta loops arranged such that the beta strands form a barrel

Topology diagram

represent elements of secondary structure and the relationships among segments of secondary structures in a protein

Protein Family/Superfamily

families are proteins with similarity in primary or tertiary structure and function; superfamilies are 2+ families with little sequence similarity but same major structural motif and function

Cellular Proteostasis

• involves numerous pathways that regulate the folding, unfolding, and degradation of proteins

• continual maintenance of the active set of cellular proteins required under a given set of conditions

Native Proteins

proteins that are synthesized, form intermediates, or chaperone assisted folding

• contain beta sheet structure which oligomerizes

Misfolded Protein

refolded or form aggregates which either leads to a disease state or degradations

Peptide Synthesis Direction

5’ → 3’ from mRNA to polypeptides to ribosomes

What influences stability of protein folding

• Thermodynamics:

  • Entropy: negative and unfavorable because folding decreases randomness

  • Enthalpy: favorable (negative) because it creates stable internal bonds

  • Free energy

  • Hydrophobic effect: increases entropy of water by shoving it out of the hydrophilic cage

• Bonding interactions

  • Hydrogen bonds

  • Ionic bonds

  • hydrophobic bonds/van der waals

  • disulphide bonds

• amino acid sequence, denaturation, and renaturation

Main driving forces for protein folding

Balance of 3 factors: negative (unfavorable) entropy, negative (favorable entropy), and hydrophobic effect (favorable)

Hydrophobic effect is the MAIN driving force for folding

Hydrophobic interactions in protein folding

• water around unfolded protein is ordered and structured

• proteins fold with hydrophobic residues on the interior

• folding removes hydrophobic and water interactions

• increases the entropy of the protein-water system because water is less ordered

Denaturation

loss of 3D structure sufficient to cause loss of function which often leads to protein precipitation

Renaturation

process by which certain denatured globular proteins regain their native structure and biological activity

What determines tertiary structure

Amino acid sequence

• Anfinsen experiment: amino acid contains also info needed to fold the protein chain

Process of polypeptide folding

Stepwise process: fast!

• local secondary structures fold first: ionic interactions and hydrogen bonding

• longer range interactions follow: hydrophobic effect

• process continues until the entire polypeptide folds

Levinthal’s paradox

mathematically impossible for protein folding to occur by randomly trying every conformation until the lowest one is found

• protein folding is hierarchal

Free Energy Funnel

unfolded states, high degree of conformational entropy, high free energy → entropy goes down in the funnel

Chaperone proteins

facilitate correct folding pathways

Hsp (heat shock proteins)

bind to hydrophobic regions and facilitate protein folding

chaperonins

required for the folding of proteins that do not fold spontaneously: assist polypeptide folding into native structure

ex) GroEL-GroES complex

Amyloid Fiber

protein secreted in a misfolded state and converted to an insoluble extracellular fluid (leads to diseases like Alzheimer’s and type 2 diabetes)

• formed by misfolded beta amyloid which promotes aggregation