Understanding a point mutation signature D54K in the caspase activation recruitment domain of NOD1 capitulating concerted immunity via atomistic simulation.

Point mutation D54K in the human N-terminal caspase recruitment domain (CARD) of nucleotide-binding oligomerization domain -1 (NOD1) abrogates an imperative downstream interaction with receptor-interacting protein kinase (RIPK2) that entails combating bacterial infections and inflammatory dysfunction. Here, we addressed the molecular details concerning conformational changes and interaction patterns (monomeric-dimeric states) of D54K by signature-based molecular dynamics simulation. Initially, the sequence analysis prioritized D54K as a pathogenic mutation, among other variants, based on a sequence signature. Since the mutation is highly conserved, we derived the distant ortholog to predict the sequence and structural similarity between native and mutant. This analysis showed the utility of 33 communal core residues associated with structural-functional preservation and variations, concurrently served to infer the cryptic hotspots Cys39, Glu53, Asp54, Glu56, Ile57, Leu74, and Lys78 determining the inter helical fold forming homodimers for putative receptor interaction. Subsequently, the atomistic simulations with free energy (MM/PB(GB)SA) calculations predicted structural alteration that takes place in the N-terminal mutant CARD where coils changed to helices (45 _{α3- L4-α4-L6- α6}83) in contrast to native (45_{T2-L4-α4-L6-T4}83). Likewise, the C-terminal helices 93_T1-α7105 connected to the loops distorted compared to native 93_α6-L7105 may result in conformational misfolding that promotes functional regulation and activation. These structural perturbations of D54K possibly destabilize the flexible adaptation of critical homotypic _NOD1CARD-CARD_RIPK2 interactions (_α4Asp42-Arg488_α5 and _α6Phe86-Lys471_α4) is consistent with earlier experimental reports. Altogether, our findings unveil the conformational plasticity of mutation-dependent immunomodulatory response and may aid in functional validation exploring clinical investigation on CARD-regulated immunotherapies to prevent systemic infection and inflammation.