Structural basis of muscarinic receptor subtypes: Importance of a single amino acid of NPxxY motif

Background and purpose: Short linear motifs are important for GPCR functions. There are differences between these motifs. Serendipity was the major role for this manuscript. Experimental approach: Regular expressions and Bash commands of linux environment including grep and multiple sequence alignment of adrenergic and muscarinic receptors of UniProt and 3D structures of PDB were used. Key results: Motif differences between adrenergic and muscarinic receptors were found. CWxP and NPxxY motifs were found on adrenoceptors but CWxP was absent on muscarinic receptors (x denotes any amino acid). NP[VIL]IY was found on adrenergic and NP[FLMAVI]CYA was found on muscarinic receptors as members of NPxxY motifs on 7th transmembrane alpha-helix. The only variable point of both motifs was at amino acids7.51 of adrenergic and muscarinic receptors. NP[FLMAVI7.51]CYA was evolutionary conserved and unique for muscarinic receptors and could not be found on any protein of any other living thing. Residues at 7.51 were found muscarinic receptor subtype specific. For example NPVCYA is unique for M3 and found only on M3 muscarinic receptors in the protein universe. Conclusion and implications: The motif NP[FLMAVI7.51]CYA and its residues at single variable point (residues7.51) are structural basis of the muscarinic receptor subtypes. New ligands interacting with this motif and acting on this fine tuning point are expected to be subtype-specific with minimal adverse reactions.