Transcript Crystal structure of the dimer

Crystal structure of the dimer
The crystal of the activated dimer shows two possible 
dimer interactions:
“Back to back”
“Head to head”
Reconstruction of the crystal
Open 1MOX.pdb in Pymol
Generate symmetry mates within 4Å (see figure).
Present all by cartoons, hide lines and color by chain
(rainbows).
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Building the head to head dimer
Display->Sequence,
mode: chains
Select the c and d chains
of each symmetry mate
to decide which one
correspond to the “head
to head” dimer. Try to
show only two cell unit
each time.
Delete the unnecessary
symmetry mates
(Actions-> delete object).
Delete the unnecessary
chains (actions-> remove
atoms).
Type in the command
line “select all” and save
molecule (sele) as a new
pdb file.
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Accessible surface area
The accessible surface area (ASA) is the surface area of a
biomolecule (protein, DNA, etc.) that is accessible to a
solvent.
The ASA is usually quoted in square ångstrom.
This algorithm uses a sphere (of solvent) of a particular
radius to 'probe' the surface of the molecule.
A typical value is 1.4Å, which approximates the radius of a
water molecule.
Another factor that affects the results is the definition of the
VDW radii of the atoms in the molecule under study
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Accessible surface area
Accessible surface area
1. Show-> as -> surface
2. Command line: set solvent_radius, 5 (default is 1.4)
Show-> as-> spheres
Calculating the interface area
Interface = ASA1 + ASA2 - ASAdimer
Calculating the interface area
Interface = ASA1 + ASA2 - ASAdimer
Calculating the interface area
http://www.bioinformatics.sussex.ac.uk/protorp/
Protorp server
Choose option 3, upload the pdb file and •
indicate which chains (protein chains),
submit.
click the results link and look for “interface •
accessible surface area (Å2).
Do the same for the other dimer. •
Calculating the interface area
“Back to back”
“Head to head”
1117Ǻ2
443Ǻ2
Calculating the interface area
“Back to back”
“Head to head”
1117Ǻ2
443Ǻ2
Biologically relevant protein-protein interfaces usually bury more than
700Ǻ2 of surface per molecule and often about 1000Ǻ2.

More reasons for choosing
the “back to back” dimer
“Back to back”
“Head to head”
Better symmetry•
Sequence conservation at the dimer interface•
Characteristic of receptors mutated at both interfaces•
Sequence Conservation
ConSurf result for 1MOX (back to back dimer): •
http://consurf.tau.ac.il/results/1227539893/output.html
Download the files for pymol and follow the instructions in •
the website.
(if you get an error when trying to run the py file try to specify the full path;
C:\...\consurf_new.py)
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Consurf result for the head to head dimer: •
http://consurf.tau.ac.il/results/1227621856/output.html
Sequence Conservation
“Back to back”
“Head to head”
hydrophobicity
Open 1MOX in Pymol
Paste this script to the command line:
Present the amino acids in the
interface between the two chains in
spheres. (use PROTORP results)
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set_color color_ile, [255,255,255]
color color_ile, resn ile
set_color color_leu, [255,255,0]
color color_leu, resn leu
set_color color_phe, [255,255,108]
color color_phe, resn phe
set_color color_val, [255,255,137]
color color_val, resn val
set_color color_ala, [255,255,235]
color color_ala, resn ala
set_color color_gly, [255,255,255]
color color_gly, resn gly
set_color color_cys, [216,255,216]
color color_cys, resn cys
set_color color_ser, [177,255,177]
color color_ser, resn ser
set_color color_thr, [147,255,147]
color color_thr, resn thr
set_color color_met, [128,255,128]
color color_met, resn met
set_color color_trp, [128,255,128]
color color_trp, resn trp
set_color color_pro, [0,252,6]
color color_pro, resn pro
set_color color_tyr, [0,230,50]
color color_tyr, resn tyr
set_color color_gln, [0,214,83]
color color_gln, resn gln
set_color color_his, [0,193,125]
color color_his, resn his
set_color color_lys, [0,184,142]
color color_lys, resn lys
set_color color_asn, [0,180,151]
color color_asn, resn asn
set_color color_glu, [0,153,205]
color color_glu, resn glu
set_color color_asp, [0,123,255]
color color_asp, resn asp
set_color color_arg, [0,0,255]
color color_arg, resn arg
Hydrophobicity
“Back to back”
“Head to head”
The “back to back” structure rules out
dimerization of the ligand!
“Back to back”
“Head to head”
The “back to back” structure rules out
dimerization of the ligand!
The receptor dimerization is mediated primarily by the interaction (1
between two EGF molecules.
The receptor dimerization is mediated primarily by the “bivalency” (2
of the ligand to the receptor.
A “receptor-mediated” mechanism, in which EGF binding induces (3
conformational changes of EGFR so as to expose its dimerization
surface.
Outline
RTKs and the ErbB family 
Ligand-induced dimerization of EGFR 
Mechanism of EGFR activation 
ErbB2 
Crystal structure of the inactive
receptor
Inactive (tethered)
Active (extended)
Crystal structure of the inactive
receptor
Dimerization
arm
Buried in
tethered
form
Inactive (tethered)
Active (extended)
The inactive receptor
The ligand binds between domains I and III, but they are not proximal in
the tethered form, so the tethered form has a lower ligand affinity.

The dimerization is mediated by domain II, but it is buried in the
tethered form, so dimerization is only possible in the extended form.

EGFR activation mechanism
The two structures give us a snapshot look at the two conformations
(tethered and extended) but they still do not explain the activation
mechanism.

One possibility is that the ligand binding actively causes the
conformation change, but there are some evidence against this.

References
Ogiso H, Ishitani R, Nureki O, Fukai S, Yamanaka M, Kim JH, Saito K, Sakamoto A,
Inoue M, Shirouzu M, Yokoyama S. Crystal structure of the complex of human
epidermal growth factor and receptor extracellular domains. Cell. 2002 Sep
20;110(6):775-87.
Burgess AW, Cho HS, Eigenbrot C, Ferguson KM, Garrett TP, Leahy DJ, Lemmon MA,
Sliwkowski MX, Ward CW, Yokoyama S. An open-and-shut case? Recent insights into
the activation of EGF/ErbB receptors. Mol Cell. 2003 Sep;12(3):541-52. Review.
Garrett TP, McKern NM, Lou M, Elleman TC, Adams TE, Lovrecz GO, Zhu HJ, Walker
F, Frenkel MJ, Hoyne PA, Jorissen RN, Nice EC, Burgess AW, Ward CW. Crystal
structure of a truncated epidermal growth factor receptor extracellular domain bound
to transforming growth factor alpha. Cell. 2002 Sep 20;110(6):763-73.
1mox.pdb