Unit II- Forces and Structures

Question 1

Van der Waal’s interactions

Answer 1

• mutually induced dipole
• atoms like to be near each other
• more or less independent of atom type
• manifested in proteins by tight packing of interior atoms
• proteins fold as to maximize van der Waals energy, evident by packed cores
• Geckos use VdW forces to adhere to a variety of surfaces
• only one packing solution, if you change the shape of one of the interior pieces that leaves a gap and causes crowding, it will destabilize the protein

Question 2

The hydrophobic effect

Answer 2

• tendency of nonpolar molecules to interact with each other rather than with water
• Nonpolar= no charges, no dipoles, no hydrogen bonding groups
• the hydrophobic effect is the major driving force for folding

Arg- water loving, 1 molecule in hexane, 300 billion in water
Leu- water hating, 1 molecule in water, 760 in hexane

-effect is due exclusively to the properties of water, clathrate-like structures are energetically costly

Question 3

Secondary structures

Answer 3

• when you turn on the noncovalent forces

- alpha-helix, beta-sheet, beta-turn

Question 4

The alpha helix

Answer 4

• most frequently observed secondary structure in proteins
• average protein about 31% alpha helix
• in the alpha helix the bonds are between the peptide NH of residue i and the peptide CO of residue i+4
• the side chains extend outward and spiral around the rod
• has periodicity of 3.6 residues
• rise 1.5 angstroms (4.5 if fully extended, therefore helix is compressed)
• helical rotation = 100 degrees per residues
• every forth amino acid is close in space

Question 5

Why 3.6 residue repeat

Answer 5

• favorable backbone dihedral angles
• near optimal hydrogen bond geometry
• good van der Waals contacts between backbone atoms
• other types of helices possible, but less common

backbone-backbone interactions
backbone-side chain interactions
side chain-side chain interactions: electrostatic (e.g. Lys-Glu), H-bonding (Asn-Asp), hydrophobic (leu-Val), and van der Waals

Question 6

Amino acid preference for alpha helix formation

Answer 6

Helix former: Ala
-lack of side chain suggests “default” backbone conformation is helical

Strong helix breaks: Pro, Gly

• Pro lacks NH to hydrogen bond
• Gly is flexible

Medium helix breakers: B-branched or bulky (Val, Thr, Trp, Phe)
-lose much rotational freedom (entropy) when in a helix

Helix indfferent: long, straight chains (Arg, Lys, Glu
-lose less rotational freedom

Question 7

Alpha helix and proteins

Answer 7

• seldom big enough to fully bury a helix in the interior
• more often one side of the helix will face towards the interior and other will face out
• stabilized by burial of nonpolar surface on the protein face and exposure of polar groups on the solvent face
• many proteins are tightly associated with cell membranes
• most membrane anchors consist of several helicecs containing mostly nonpolar residues
• these residues strongly interact with the hydrocarbon interior of the lipid bilayer via the hydrophobic effect and generally do not dissociate unless the membrane is disrupted by detergents

Question 8

The beta sheet

Answer 8

• the average protein contains 28 % beta sheet, the second most common type of secondary structure
• made up of two or more beta-strands
• the polypeptide chain is straight and nearly completely extended
• H -bonds are formed between peptide groups of each strand
• Beta sheets are fully hydrogen bonded structures
• pleated appearance
• pointing in the same direction (parallel) less common because H bonds are slightly bent, or head to tail (antiparallel)

Question 9

Beta sheet info

Answer 9

• favorable backbone dihedral angles
• phi, psi angles less than 140 degrees, chain is nearly fully extended
• rise 3.5 angstrom
• periodicity=2
• straight hydrogen bonds between strands
• interaction between adjacent strands (tertiary interaction) - Amyloid
• tend to form insoluble aggregates
• amphipathic

Question 10

The reverse Beta turn

Answer 10

• 1/4 of protein structure
• several types
• much sequence variability
• required: Gly
• preferred: Pro
• other positions: solvent exposed, polar residues

Question 11

Irregular structure

Answer 11

• sometimes called “random coil”
• generally only random in the sense that it is not periodic
• usually has specific structure
• surface loops are critical to function
• loops help give proteins their individuality

Question 12

Motifs and domains

Answer 12

• secondary structures are assembled in a limited number of patterns to form domains
• domains are added, deleted, and swapped to generate new structures and functions
• Motifs- small, simple structures, unstable
• Domains-visual thing, bigger more complex

Question 13

Helix-turn-helix Motif

Answer 13

• consist of short stretches of secondary structure and are usually not stable by themselves
• used in molecular recognition-to bind other molecules tightly and specifically
• one of most common elements that recognizes specific sequences of DNA
• -found in homeodomains
• consists of a recognition alpha-helix and a support alpha-helix, connected by turn
• sits on the major groove and binds to specific sequence of polypeptide

-the i,i+3, i+4 spacing of hydrophobic residues helps determine that the polypeptide chain adopts the HTH fold

Question 14

Zinc finger motif

Answer 14

• DNA binding motif
• bind DNA weakly
• TF proteins use 2-40 of these repeats to bind DNA
• Zn2+ is to help hold the structure together without an extensive hydrophobic core.
• hydrophobic side chains are still used to stabilize the helix which binds the major groove of DNA

Question 15

Coiled-coil domain

Answer 15

• domains are stable, semi-independent units of structures
• residues within the domain interact more with each other than with residues outside of it
• found in fibrous proteins and transcription factors
• heptad repeat, a-f. Hydrophobic- a and d, e and g are usually opposite charge

-also used in protein-protein recognition, mechanical force transduction (myosin tails), and viral penetration

Question 16

DNA binding domain of GCN4 TF

Answer 16

• DNA binding domain of GCN4 transcription factor is a parallel, coiled-coil homodimer
• the C terminal half of the molecule is a classical heptad repeat “leucine zipper”
• in absence of DNA, the DNA binding half of GCN4 is not structured, the extensive interactions with DNA (electrostatic, H-bonding, hydrophobic, van der Waals) is what causes the N-terminal half to adopt the fully helical structure shown

Question 17

Influenza haemmagglutin

Answer 17

• trimer of identical subunits, made of two domains
• the stalk domain is comprised of a triple-stranded coiled coil with each monomer contributing one helix
• it is a molecular harpoon, binds to human cells via the head domain
• virus is endocytosed into the cell and in vesicles below -5.0, a conformational change take place in which an extended loop folds into an alpha helix
• the coiled coil can then length and then it can get out of the vacuolar membrane, release viral RNA into the cell

Question 18

Structure based design of viral entry inhibitors

Answer 18

• many viruses in addition to influenza use similar harpooning mechanisms to gain entry into cells (HIV, SARS, ebola)
• the mechanism of entry involves membrane distortion mediated by the harpoon protein
• small molecule inhibitors can be developed that specifically target different steps in this mechanism

HIV gets in via gp120 and gp41 binding CD4 and CXCR4/CCR5 coreceptors, harpooning because of receptor
-FDA fusion inhibitors

Question 19

Gp41 bound to a mirror image peptide inhibitor

Answer 19

• D-amino acids; L-amino acids are the normal enanatiomer
• D version of the peptide will bind to the L version of gp41 with exactly the same affinity and specificity
• the D peptide inhibitor above binds to gp 41 more tightly than Fuzeon and is not degraded by body proteins

Question 20

Bacterial pore-forming toxin

Answer 20

• how hydrophobic/ hydrophilic amino acid patterning determines structure-function relationships
• how different representations of protein structure are useful to visualize structure-function relationships
• one mechanism by which pathogenic microorganisms cause sickness and death
• how small molecules may be devised to combat these microorganisms
• pore forming proteins self associate to and insert into the plasma membrane of the host cells, forming channels of up to several nanometers
• this effectively pokes a hole in the cell membrane, causing leakage and cell death

Question 21

Solvent accessible surface area of hemolysin

Answer 21

• gives the most accurate view of how a protein actually looks in the cell or in solution
• the outside of the beta barrel domain interacts with the plasma membrane- is completely devoid of charges and is exclusively and is hydrophobic
• the regions of the protein that interact with aqueous solution are speckled with charages

Question 22

Proteins going wrong

Answer 22

• when something goes wrong in the body, a protein defect is almost always to blame
• protein folding in virto and in vivo
• no cellular factors required
• native structures is lowest energy state
• 3D structure is encoded in 1D sequence

Question 23

Protein misfolding

Answer 23

• clear that is a complex process which involves formation of intermediates
• many causes folding follows a sequential mechanism
• the protein is vulnerable to aggregation when in the partially folded formations because they tend to expose hydrophobic residues that have yet to be buried in the hydrophobic core

-the cell is extremely crowded- competition between hydrophobic collapse within the same molecule and with other protein and membrane molecules (aggregate formation)