1. vFLIP-IKKg Blocker Edith Chan WIBR

2. IKKg NF-kB Pathway • The Nf-kB pathway is related to inflammatory responses, cell death, and oncogenesis. • Kaposi’s sarcoma herpesvirus (KSHV) encoded FLIP (vFLIP) binds to IKKg to activate NFkB. • Similarly, in human, cFLIP can associated with IKKg and thus has a wider application in cancer therapies.

3. Work Study • The study focuses on using X-ray crystal structures, biophysical screening and structure based design to identify blockers of the vFLIP-IKKg and p22-cFLIP-IKKg interaction, conducting lead optimisation and identifying a development candidate. • My immediate actions are • Understanding of the interaction of the vFLIP-IKKg complex using X-ray crystal structure • Selection of compounds mimicking the IKKg interaction of the complex

4. vFLIP C-terminus IKKg N-terminus E B A D X-ray crystal structure • Our collaborators at BBK have solved the structure between vFLIP-IKKg (3cl3). • Full length IKKg 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-IKKg complex. • The two vFLIP molecules come together solely through interactions between the two IKKg chains.

5. Protein-Protein Interactions • Each of the IKKg 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 IKKg)

6. 226 246 193QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE253 Peptide tool compound vFLIP • Full length IKKg 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 IKKg 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 IKKg be sufficient to bind with vFLIP? • Can a shorter peptide adopt helical conformation on its own or by recognition to vFLIP? C-terminus N-terminus IKKg

7. 226 238 246 193QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE253 1 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE 2 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE 3 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE 4 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE 5 QQHSVQVDQLRMQGQSVEAALRMERQAASEEKRKLAQLQVAYHQLFQEYDNHIKSSVVGSE Proposed tool Peptides LengthAA spin • Longer peptide include C-terminus 30aa aa224-253 • Spin the entire interaction with vFLIP 21aa aa226-246 • Peptide that covers cleft1 interaction 16aa aa231-246 • Peptide that covers cleft2 interaction 15aa aa226-240 • Peptide that covers all essential interaction 12aa aa231-242