Molecular Biophysics
12824 BCHS 6297
Lecturers held Tuesday and Thursday 10 AM – 12 Noon 402B-HSC
Aims of Course
• Understand role of physics in protein structure
• Overview of the Electronic Spectroscopies
• Understand the application of kinetics and thermodynamics to study enzyme catalysis and protein folding
• Basics of NMR and X-ray crystallography
Suggested texts
• Principles of Physical Biochemistry, van Holde • Structure and Mechanism in Protein Science,
Fersht• On-line resourses • Understanding NMR spectroscopy, by James
Keelerhttp://www-keeler.ch.cam.ac.uk/lectures/• Principles of Protein Structure Using the Internet,http://www.cryst.bbk.ac.uk/PPS2/course/index.html
Structural Biology of the HIV proteome
Molecular Forces in Protein Structure
• Interactions, forces and energies • Covalent Interactions • Non-bonded Interactions • Electrostatic interactions: salt bridges, hydrogen
bonds, partial charges and induction • The Lennard-Jones potential and van der Waals
Radii • The effect of solvent and hydrophobic interactions • Dielectric effects • The hydrophobic effect
Covalent bondThe force holding two atoms together by the sharing of a pair of electrons.
H + H H:H or H-H
The force: Attraction between two positively charged nuclei and a pair of negatively charged electrons.
Orbital: a space where electrons move around.
Electron can act as a wave, with a frequency, and putting a standing wave around a sphere yields only discrete areas by which the wave will be in phase all around. i.e different orbitals.
Polarity of Bonds H | + -
CH3OH H—C—OH C O | H
or even stronger polarity
H + - + -
H C O C O
O> N> C, H electronegativity - + + - + -
O H C N C O
Geometry also determines polarity
• + - • while C Cl is polar
carbon tetrachloride is not. The sum of the vectors equals zero and it is therefore a nonpolar molecule
CCl4 = 1+2+3+4 = 0
CCl Cl
Cl Cl1
2
3
4
CCl Cl
Cl
2
3
4 H
CHCl3 is polar
Electrostatic interactions
by coulombs law F= kq1q2 q are charges r2D r is radius
D = dielectric of the media, a shielding of charge. And k =8.99 x109Jm/C2
D = 1 in a vacuumD = 2-3 in greaseD = 80 in water
Responsible for ionic bonds, salt bridges or ion pairs,optimal electrostatic attraction is 2.8Å
Dielectric effect Dhexane 1.9benzene 2.3diethyl ether 4.3CHCl3 5.1acetone 21.4Ethanol 24methanol 33H2O 80HCN 116
H2O is an excellent solvent and dissolves a large array of polar molecules.
However, it also weakens ionic and hydrogen bondsTherefore, biological systems sometimes exclude H2O to
form maximal strength bonds!!
Hydrogen bonds
O-H N N-H O 2.88 Å 3.04 Å
H bond donor or an H bond acceptor
N H O C
3-7 kcal/mole or 12-28 kJ/molevery strong angle dependence
A hydrogen bond between two water molecules
.
van der Waals attraction
Non-specific attractions 3-4 Å in distance (dipole-dipole attractions)
Contact Distance
ÅH 1.2 1.0 kcal/molC 2.0 4.1 kJ/molN 1.5 weak interactionsO 1.4 important when many atomsS 1.85 come in contactP 1.9
Can only happen if shapes of molecules match
Hydrophobic interactionsNon-polar groups cluster together
G = H - TS
The most important parameter for determining a biomolecule’s shape!!! Entropy order-disorder. Nature prefers to maximize entropy “maximum disorder”.
Enthalpy How can structures form if they are unstable?
Structures are driven by the molecular interactions of the water!
STRUCTURED WATER
A cage of water molecules surrounding the non-polar molecule
This cage has more structure than the surrounding bulk media.
G = H -TS
Entropy decreases!! Not favorable! Nature needs to be more disorganized. A driving force.
SO
To minimize the structure of water the hydrophobic molecules cluster together minimizing the surface area. Thus water is
more disordered but as a consequence the hydrophobic molecules become ordered!!!
Proton and hydroxide mobility is large compared to other ions
• H3O+ : 362.4 x 10-5 cm2•V-1•s-1
• Na+: 51.9 x 10-5
• Hydronium ion migration; hops by switching partners at 1012 per second.
Free energy of transfer for hydrocarbons form water to organic solvent
CH4 in H2O CH4 in C6H6 11.7 -22.6 -10.9
CH4 in H2O CH4 in CCl4 10.5 -22.6 -12.1
C2H6 in H2O C2H6 in C6H6 9.2 -25.1 -15.9
Process H -TS G
Amphiphiles form micelles, membrane bilayes and vesicles
• A single amphiphile is surrounded by water, which forms structured “cage” water. To minimize the highly ordered state of water the amphiphile is forced into a structure to maximize entropy
G = H -TS driven by TS
Amino Acids:The building blocks of proteins
amino acids because of the carboxylic and amino groupspK1 and pK2 respectively pKR is for R group pK’s
pK1 2.2 while pK2 9.4
pK1pK2
In the physiological pH range, both carboxylic and amino groups are completely ionized
Amino acids are Ampholytes
They can act as either an acid or a base
They are Zwitterions or molecules that have both a positive and a negative charge
Because of their ionic nature they have extremely high melting temperatures
Amino acids can form peptide bonds
Amino acid residue
peptide units
dipeptides
tripeptides
oligopeptides
polypeptides
Proteins are molecules that consist of one or more polypeptide chains
Peptides are linear polymers that range from 8 to 4000 amino acid residues
There are twenty (20) different naturally occurring amino acids
Characteristics of Amino AcidsCharacteristics of Amino AcidsThere are three main physical categories to describe amino acids:
1) Non polar “hydrophobic” nine in allGlycine, Alanine, Valine, Leucine, Isoleucine,
Methionine, Proline, Phenylalanine and Tryptophan
2) Uncharged polar, six in allSerine, Threonine, Asparagine, Glutamine Tyrosine,
Cysteine
3) Charged polar, five in allLysine, Arginine, Glutamic acid, Aspartic acid, and
Histidine
Amino Acids
You must know:
Their namesTheir structureTheir three letter codeTheir one letter code
H2N CH C
CH2
OH
O
OH
Tyrosine, Tyr, Y, aromatic, hydroxyl
Cystine consists of two disulfide-linked cysteine residues
Acid - Base properties of amino acids
[HA]
][Alog pK pH
-
ji pKpK2
1 pI
Isoelectric point: the pH where a protein carries no net electrical charge
For a mono amino-mono carboxylic
residue pKi = pK1 and pKj = pK2 ; for
D and E, pKi = pK1 and pKj - pKR ;
For R, H and K, pKi = KR and pKj =
pK2
The tetra peptide Ala-Tyr-Asp-Gly or AYDG
Greek lettering used to identify atoms in lysine or glutamate
Optical activity - The ability to rotate plane - polarized light
Asymmetric carbon atom
Chirality - Not superimposable
Mirror image - enantiomers
(+) Dextrorotatory - right - clockwise
(-) Levorotatory - left counterclockwise
Na D Line passed through polarizing filters.
ionconcentratlength x path
(degrees)roration observed ][ 25
D
}Operational
definition only cannot predict
absolute configurations
The Fischer Convention
Absolute configuration about an asymmetric carbon
related to glyceraldehyde
(+) = D-Glyceraldehyde
(-) = L-Glyceraldehyde
In the Fischer projection all bonds in the horizontal direction is coming out of the plane if the paper,
while the vertical bonds project behind the plane of the paper
All naturally occurring amino acids that make up proteins are in the L conformation
The CORN method for L isomers: put the hydrogen towards you and read off CO R N clockwise around the C This works for all amino acids.
An example of an amino acid with two asymmetric carbons
An example of an amino acid with two asymmetric carbons
Structural hierarchy in proteins
Color conventions
Protein Geometry
CORN LAW amino acid with L configuration
Greek alphabet
The Polypeptide Chain
Polypeptide geometry
• Pauling and Corey
Peptide bond
• C-N bond displays partial double bond character
Peptide bonds generally adopt a trans configuration
Peptide Torsion Angles
Torsion angles determine flexibility of backbone structure
Steric hindrance limits backbone flexibility
Rammachandran plot for L amino acids
Indicates energetically favorable / backbone rotamers
Regular Secondary Structure Pauling and Corey
Helix Sheet
alpha helix
Properties of the helix
• 3.6 amino acids per turn
• Pitch of 5.4 Å
• O(i) to N(i+4) hydrogen bonding
• Helix dipole
• Negative and angles,
• Typically = -60 º and = -50 º
Distortions of alpha-helices
• The packing of buried helices against other secondary structure elements in the core of the protein.
• Proline residues induce distortions of around 20 degrees in the direction of the helix axis. (causes two H-bonds in the helix to be broken)
• Solvent. Exposed helices are often bent away from the solvent region. This is because the exposed C=O groups tend to point towards solvent to maximize their H-bonding capacity
Top view along helix axis
Helical bundle
310 helix
• Three residues per turn
• O(i) to N(i+3) hydrogen bonding
• Less stable & favorable sidechain packing
• Short & often found at the end of helices
Helical propensity
Peptide helicity prediction
• AGADIR
http://www.embl-heidelberg.de/Services/serrano/agadir/agadir-start.html
Agadir predicts the helical behaviour of monomeric peptides
It only considers short range interactions
beta () sheet
• Extended zig-zag
conformation
• Axial distance 3.5 Å
• 2 residues per repeat
• 7 Å pitch
Antiparallel beta sheet
Antiparallel beta sheet side view
Parallel beta sheet
Parallel, Antiparallel and Mixed Beta-Sheets
Beta sheets are twisted
• Parallel sheets are less twisted than antiparallel and are always buried. • In contrast, antiparallel sheets can withstand greater distortions (twisting and beta-bulges) and greater exposure to solvent.