Stimulated Toll-like receptors (TLRs) trigger the signaling pathway and the first event — the recruitments and interactions of various signaling pathway molecules— occurs to defend our body from the invading pathogens. Among these adaptor proteins, MyD88 adaptor-like (MAL) plays a crucial role in mediating and interaction between the Toll/Interleukin-1 Receptor (TIR) domain-containing signaling molecules such as TLR2/4-TIR and MyD88-TIR. Because of the highly conserved amino acid sequence between the TIR domains, we designed a novel decoy peptide derived from MAL-αC helix that interacts with the TIR domain of MyD88 and TLRs. The designed peptide, MAL/MyD88-inhibitory peptide (MIP), was predicted to interact with MAL, MyD88-TIR, and TLR4-TIR domains. To facilitate cellular uptake, cell-penetrating peptides (CPPs) were linked to the N-terminal of MIP. Only MIP2, linked to penetratin (RQIKIWFQNRRMKWKK), substantially suppressed the NF-κB activation, MyD88- and TRIF-dependent TLRs signaling except TLR3. Through the SPR analysis and immunofluorescence, it is investigated that MIP2 binds to MAL, MyD88-TIR, and TLR4-TIR. Collectively, we report that our decoy peptide, MIP2, has a wide range of TLR inhibitory effects and would be effective in easing autoimmune disease symptoms.
