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vFLIP-IKK Blocker

Edith Chan

WIBR

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NF-B Pathway• The Nf-B pathway is related to

inflammatory responses, cell death, and oncogenesis.

• Kaposi’s sarcoma herpesvirus (KSHV) encoded FLIP (vFLIP) binds to IKK to activate NFB.

• Similarly, in human, cFLIP can associated with IKK and thus has a wider application in cancer therapies.

IKK

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Work Study

• The study focuses on using X-ray crystal structures, biophysical screening and structure based design to identify blockers of the vFLIP-IKK and p22-cFLIP-IKK interaction, conducting lead optimisation and identifying a development candidate.

• My immediate actions are

– Understanding of the interaction of the vFLIP-IKKcomplex using X-ray crystal structure

– Selection of compounds mimicking the IKK interaction of the complex

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X-ray crystal structure• Our collaborators at BBK have

solved the structure between vFLIP-IKK (3cl3).

• Full length IKK is 419aa long mulitdomain protein

• Both proteins are truncated– ks-vFLIP (aa1-178) [188aa]– IKKg (aa150-272) [419aa]

• The X-ray structure comprised of a dimer of two ks-vFLIP-IKKcomplex.

• The two vFLIP molecules come together solely through interactions between the two IKK chains.

IKK

vFLIPC-terminus

N-terminus

A B

D

E

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Protein-Protein Interactions

• Each of the IKK helix is interacting with a copy of the vFLIP via two adjacent vertical clefts (cleft 1 and 2)

• Cleft 1 involved more interactions between the complex, the hottest spot seems to be around Phe238 (of IKK)

6193QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE253

226 246

Peptide tool compound• Full length IKK is 419aa long mulitdomain protein. In

the X-ray structure, only the HLX2 (helix) domains (aa193-253) are co-crystallized with ks-vFLIP.

• As seen from the picture, only a portion of the IKK is interacting with vFLIP. They are about 20aa long as highlighted in a pink box.

• Our first step is to use of peptide as proof of concept tool in further experiments

• Can a shorter peptide of IKK be sufficient to bind with vFLIP?

• Can a shorter peptide adopt helical conformation on its own or by recognition to vFLIP?

IKK

vFLIP

N-terminus

C-terminus

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Proposed tool Peptides

Length AA spin1. Longer peptide include C-terminus 30aa aa224-253

2. Spin the entire interaction with vFLIP 21aa aa226-246

3. Peptide that covers cleft1 interaction 16aa aa231-246

4. Peptide that covers cleft2 interaction 15aa aa226-240

5. Peptide that covers all essential interaction 12aa aa231-242

193QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE2531 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE2 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE3 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE4 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE 5 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE

226 238 246