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The wild-type human CCK1R and CCK2R with C-terminal 15-residue depleted were subcloned into pFastBac vector with an N-terminal FLAG-tag and C-terminal 10× His-tag. A dominant-negative human Gαs (DNGαs) was generated by site-directed mutagenesis as previously described to limit G protein dissociation. Twenty-nine amino acids at the N-terminus of wild-type Gαq were replaced by the corresponding sequence in Gαi1 to facilitate the binding of scFv16. The constructs were cloned into both pcDNA3.1 and pFastBac vectors for functional assays in mammalian cells and protein expression in insect cells, respectively. All modifications of the receptor had no effect on ligand binding and receptor activation.

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CCK peptides release insulin and glucagon more potently in man and pig than in dog and rat (51, 85–87). The difference is partly due to neurons in pancreatic islets that release CCK-8 and CCK-5 in man and pig (51), whereas rat and dog islets have no such innervation (50, 51). Moreover, islet cells in man and pig also express the CCK2 receptor abundantly (73), whereas rat islet cells express mainly the CCK1 receptor (88). While the interest in the effect of CCK on the exocrine pancreas was for many years restricted to enzyme secretion, it is now well established that CCK also stimulates fluid and bicarbonate secretion.

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Cholecystokinin (CCK) receptor family belongs to class-A sevenfold transmembrane G protein-coupled receptors, and is divided into CCK1 receptor (CCK1R) and CCK2 receptor (CCK2R)1. CCK1R is mainly distributed in the gastrointestinal tract2,3,4, peripheral nervous system5,6, and some regions of the brain, e.g., the area postrema, the nucleus tractus solitarius, and the hypothalamus7,8,9,10. Hence, CCK receptors (CCKRs) regulate a variety of physiological functions including digestion, satiety, emotion regulation, pain sensation, and memory process18,19,20,21,22,23,24,25. Besides, CCKRs are expressed in the brain–gut axis which is a region critical for the transmission of information between gut and brain including satiation signals. For these functions, there are already some specific drugs designed for CCK1R, such as ceruletide, or for CCK2R, including proglumide and pentagastrin. Although all of them have been used for gastrointestinal diseases, they and some drugs under clinical trial also have the potential to treat CNS diseases, including pain and anxiety26,27.

C Superposition of SR activated CCK1R (pink) with active β2AR (dark green) and inactive β2AR (gray). D–g Close-up views of conformational changes in crucial residues involved in CCK1R (pink) and β2AR (dark green) activation. For those who prefer quick and anonymous transactions, Ck444 accepts popular cryptocurrencies like Bitcoin, Ethereum, and Litecoin. Rise to the top of the leaderboard and claim your share of the prize pool with the Ck444 Leaderboard reward.

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CCK-8 predominates in the brain, and gastrin-17 accounts for 90% of gastrin in the human body, inducing acid release30. Notably, posttranslational modifications (PTMs) are critical for CCK and gastrin activity, including sulfation of the tyrosine (TYS) at position 7 from the C-terminus in CCK and at position 6 in gastrin (Fig. 1a). Intriguingly, CCK2R binds sulfated and non-sulfated ligands equally well, whereas CCK1R exclusively responds to sulfated CCK.