Upload
race
View
33
Download
0
Tags:
Embed Size (px)
DESCRIPTION
vFLIP vs cFLIP Cleft 1. Edith Chan WIBR. cFLIP – sequence and structure. One of our objectives in this project is to find binders for the p22-cFLIP IKK g interaction. - PowerPoint PPT Presentation
Citation preview
1
vFLIP vs cFLIPCleft 1
Edith Chan
WIBR
2
cFLIP – sequence and structure• One of our objectives in this project is to find binders for the p22-
cFLIP IKK interaction.
• Sequence identity between the vFLIP and cFLIP is low, about 30%. However, p22-cFLIP has been shown to be a strong activator for NF-B.
• cFLIP homology model was built using Modeler and this 3D structure has confirmed that the two binding clefts can be clearly identified and the mutation of residues can be accommodated easily within the structure.
Green – conserved residuesCyan – binding sitesMagenta – insertion/deletion of MC159
3
vFLIP – Clef1 review• SURFNET locates two clefts on the protein surface - Cleft1 and Cleft2.
• In Cleft1, mainly F238, D242, and K246 from IKK interact with vFLIP, with F238 reaching the deepest pocket.
• In Cleft2, Q236 and E240 are pointing into the pocket. Binding site relatively more shadow.
• Cleft1 is relatively deeper than Cleft2, making it a better target for molecular design.
Cleft1Cleft2
4
vFLIP vs cFLIP - Cleft1 sequence• Below shown the residues that are involved in interaction with IKK.
• Two residues are conserved – A57 and H83.
• Mutation study on vFLIP
– A57L has impaired the forming of the complex and the activation of IKK.
– P54G shows a reduction in affinity
– Y90F retains affinity
• Mutation study on IKK– D242R mutant has rendered IKK incapable of forming a complex.
53 57 79 83 87 90 Mutation G L FvFLIP: FP--A---------------------FL--H---TMSYcFLIP: VG--A---------------------AV--H---NPHL 52 56 78 82 86 89
Mutation done on vFLIPA-L impaired effectP-G reduction in affinityY-F retain affinity
5
Size of Cleft1• The proposed Cleft1 binding site of cFLIP is larger than vFLIP, especially in two areas.
• The area around where F238 binds is much deeper in cFLIP
– L80(v) to V79(c): Leu sidechain is larger than Val.
• Similarly, a slightly larger pocket around where D242/K246 binds, although somewhat near the surface of the binding site
– S89(v) to H88(c) and Y90(v) to L89(c)
• Both of these may imply cFLIP can accommodate bigger groups.
Yellow cFLIpRed vFLIP
L80(v)V79(c)
S89(v)H88(c)
Y90(v)L89(c)
F238
6
Pharmacophore – HA• In both proteins, HA interactions (e.g. CO2-, C=O) are localized around where
Asp242 binds due to His82/Y90 or His82/Asn86.
• Since His, Tyr, and Asn are also capable of HD and HA, HD pharmacophore is also detected at that region, although the contribution is much smaller at cFLIP.
Red sphere - HA and negative charge, Blue sphere - HD and positive charge
vFLIP cFLIP
M88 T87 H83
Y90
G53
P87 N86 H82
P54
L89
7
Pharmacophore - HD• In vFLIP, HD interactions (N-H, sp3 N) can be found in two regions:
– lys246 binds (C=O from Met88)
– Near pro54 (C=O) - small probe
• In cFLIP, HD interaction is near Gly53 (from C=O) – small probe
• The loss of a HD contribution around where lys246 binds is due to a change from M88(vFLIP) to P87(cFLIP). The backbond C=O is no longer available from Pro87.
Red sphere - HA and negative charge, Blue sphere - HD and positive charge
vFLIP cFLIP
M88 T87 H83
Y90
G53
P87 N86 H82
P54
L89
8
Pharmacophore - Hydrophobic• sp3 C (hydrophobic/lipophlic interaction) and sp2 C (aromatic interaction) are all favourable in both proteins.
• However, aromatic is slightly favourable near where Phe binds at vFLIP (due to aromatic residues), while hydrophobic is slightly favourable in the cFLIP binding site.
Green sphere – sp3 type C, Purple sphere – sp2 type C aromatic
vFLIP cFLIP
M88 T87 H83 L80 F79
Y90
P87 N86 H82 V79 A78
P54 F53 G53 V52
A56
A57
L89
9
Cleft 1 - SummaryIKK- vFLIP cFLIP
F238 F53, P54, A57, F79, L80.
Aromatic/hydrophobic
V52, G53, A56, A78, V79.
Hydrophobic/aromatic
Pocket is deeper
D242
K246
H83, T87, M88, S89, Y90.
Hydrophobic
H83, N86, P87, H88, L89
Aromatic/Hydrophobic
Pocket is larger near the surface
D246 HA - H83 (sidechains) and Y90 (-OH)
HA - H82 and N86 (sidechains) L89 – no Hbond capability
K246 HD - M88 (C=O) NO HD, P87 (C=O) point away from binding site
10
Compounds to buy
• Aryl sulfonamides
• Isoindolinones – have not found suitable compounds to buy yet (ChemDiv)
• Nutlins – wait for more compounds to be ordered at the same time
11
Sulfonamides
S
N+
NHO
O
O
O
S
N+
N
O
O
O
O
S
N
O
O
O
OH
S
N
O
O
O
OH
Cl
S NH
O
O
O
OH
Cl
SN
O
O
O
OH
SN
OO
OOH
O
SNH
OO
O
NH2
O
S
NH
O O
O
S
N
O
O
O
NH2O
• ChemBridge
• 1mg or 5mg - $38
• 10 compounds selected.