Upload
alexis-henderson
View
216
Download
0
Embed Size (px)
Citation preview
Protein Folding & BiospectroscopyProtein Folding & Biospectroscopy
F14PFB
David Robinson
Mark Searle
Jon McMasterhttp://robinson.chem.nottingham.ac.uk/teaching
Module OverviewModule OverviewThe course will develop an understanding of protein
structure, stability, design and methods of structural analysis; understand the protein folding problem and experimental approaches to the analysis of protein folding kinetics and the application of site-directed mutagenesis. A range of experimental spectroscopic techniques will be introduced to probe protein structure and stability based on secondary structure and tertiary interactions and to probe the nature of the active site of metalloproteins using equilibrium and time-resolved spectroscopy.
Protein FoldingProtein Folding
1. Introduction
2. Protein Structure
3. Interactions
4. Protein Folding Models
5. Biomolecular Modelling
6. Bioinformatics
Handouts: http://robinson.chem.nottingham.ac.uk/teaching/F14PFB
3D Structure of Myoglobin3D Structure of Myoglobin
- first to be determined by x-ray crystallography
- revealed how the protein bound heme (loaded with oxygen) and gave the first detailed look at a protein structure
- now 10,000’s of protein structures are known
The many functions of proteinsThe many functions of proteins
Mechanoenzymes: myosin, actin Rhodopsin: allows vision Globins: transport oxygen Antibodies: immune system Enzymes: pepsin, renin, carboxypeptidase A Receptors: transmit messages through
membranes• And hundreds of thousands more…
Proteins are chains of amino acidsProteins are chains of amino acids Polymer – a molecule composed of repeating units
Amino acid compositionAmino acid composition Basic Amino Acid
Structure:• The side chain, R,
varies for each ofthe 20 amino acids
C
RR
C
H
NO
OHH
H
Aminogroup
Carboxylgroup
Side chain
The Peptide BondThe Peptide Bond
Dehydration synthesis Repeating backbone: N–C –C –N–C –C
• Convention – start at amino terminus and proceed to carboxy terminus
O O
Peptidyl polymersPeptidyl polymers A few amino acids in a chain are called a
polypeptide. A protein is usually composed of 50 to 400+ amino acids.
Since part of the amino acid is lost during dehydration synthesis, we call the units of a protein amino acid residues.carbonylcarbonylcarboncarbon
amideamidenitrogennitrogen
Side chain propertiesSide chain properties Recall that the electronegativity of carbon is at
about the middle of the scale for light elements• Carbon does not make hydrogen bonds with water
easily – hydrophobic• O and N are generally more likely than C to h-bond
to water – hydrophilic We group the amino acids into three general
groups:• Hydrophobic• Charged (positive/basic & negative/acidic)• Polar
The Hydrophobic Amino AcidsThe Hydrophobic Amino Acids
Proline severelyProline severelylimits allowablelimits allowableconformations!conformations!
The Charged Amino AcidsThe Charged Amino Acids
The Polar Amino AcidsThe Polar Amino Acids
More Polar Amino AcidsMore Polar Amino Acids
And then there’s…And then there’s…
Amino acids
Planarity of the peptide bondPlanarity of the peptide bond
Phi () – the angle of rotation about the N-C bond.
Psi () – the angle of rotation about the C-C bond.
The planar bond angles and bond lengths are fixed.
Phi and psiPhi and psi = = 180° is
extended conformation
: C to N–H : C=O to C
C
C=O
N–H
The Ramachandran PlotThe Ramachandran Plot
G. N. Ramachandran – first calculations of sterically allowed regions of phi and psi
Note the structural importance of glycine
Observed(non-glycine)
Observed(glycine)Calculated
Four levels of protein structureFour levels of protein structurePrimary: amino acid sequence
Ser Val Tyr Cys
Four levels of protein structureFour levels of protein structure
Primary: amino acid sequence
Secondary: regular, repeated coiling
and folding of
polypeptide backbone
Four levels of protein structureFour levels of protein structure
Primary: amino acid sequence
Secondary: regular, repeated coiling and folding of polypeptide backbone
Tertiary: complete three-dimensional structure
Quaternary: arrangement of
subunits (in multisubunit
protein)
Secondary structureSecondary structure• Regular, repeated coiling and folding of
polypeptide backbone Due to hydrogen bonding Two patterns
(alpha) helix (beta) sheet
TertiaryTertiary
Complete three-dimensional structure Due to weak interactions between side (R)
groups as well as covalent disulfide bonds
Weak interactionsHydrogen bondsElectrostatic interactions (ionic bonds)Hydrophobic interactionsVan der Waals interactions
Tertiary Tertiary structure structure
formed through formed through side chain side chain
interactionsinteractions
TertiaryTertiary
Complete three-dimensional structure
Composed of:
• Motifs: specific combinations of secondary structural elements
• Domains: structurally independent units
MotifsMotifs
specific combinations of secondary structural elements
DomainsDomains
Structurally
independent units
Two different
binding domains
to bind two different
molecules
TertiaryTertiary
Complete three-dimensional structure
Native conformation: functional structure
Most stable conformation
TertiaryTertiaryFibrous Proteins = extended filaments
or
Globular proteins =
compact folded structure