We mutated key amino acids of the human variant of the M1 muscarinic receptor that target ligand binding, receptor activation, and receptor-G protein interaction. We compared the effects of these mutations on the action of two atypical M1 functionally preferring agonists (N-desmethylclozapine and xanomeline) and two classical non-selective orthosteric agonists (carbachol and oxotremorine). Mutations of D105 in the orthosteric binding site and mutation of D99 located out of the orthosteric binding site decreased affinity of all tested agonists that was translated as a decrease in potency in accumulation of inositol phosphates and intracellular calcium mobilization. Mutation of D105 decreased the potency of the atypical agonist xanomeline more than that of the classical agonists carbachol and oxotremorine. Mutation of the residues involved in receptor activation (D71) and coupling to G-proteins (R123) completely abolished the functional responses to both classical and atypical agonists. Our data show that both classical and atypical agonists activate hM1 receptors by the same molecular switch that involves D71 in the second transmembrane helix. The principal difference among the studied agonists is rather in the way they interact with D105 in the orthosteric binding site. Furthermore, our data demonstrate a key role of D105 in xanomeline wash-resistant binding and persistent activation of hM1 by wash-resistant xanomeline.

• MeSH
• agonisté muskarinových receptorů farmakologie   MeSH
• buněčná membrána účinky léků   MeSH
• CHO buňky MeSH
• Cricetulus MeSH
• inositolfosfáty metabolismus   MeSH
• karbachol farmakologie   MeSH
• křečci praví MeSH
• molekulární modely MeSH
• mutace MeSH
• oxotremorin farmakologie   MeSH
• pyridiny farmakologie   MeSH
• receptor muskarinový M1 agonisté   genetika   MeSH
• receptory spřažené s G-proteiny účinky léků   MeSH
• thiadiazoly farmakologie   MeSH
• vápník metabolismus   MeSH
• vazebná místa účinky léků   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH