Dopamine D 2 receptor agonist binding kinetics-role of a conserved serine residue

Ågren, Richard; Stepniewski, Tomasz Maciej, 1988-; Zeberg, Hugo; Selent, Jana; Sahlholm, Kristoffer

doi:http://dx.doi.org/10.3390/ijms22084078

Dopamine D 2 receptor agonist binding kinetics-role of a conserved serine residue

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Ågren R, Stepniewski TM, Zeberg H, Selent J, Sahlholm K. Dopamine D 2 receptor agonist binding kinetics-role of a conserved serine residue. Int J Mol Sci. 2021;22(8):4078. DOI: 10.3390/ijms22084078

Abstract

The forward (kon) and reverse (koff) rate constants of drug-target interactions have important implications for therapeutic efficacy. Hence, time-resolved assays capable of measuring these binding rate constants may be informative to drug discovery efforts. Here, we used an ion channel activation assay to estimate the kons and koffs of four dopamine D2 receptor (D2R) agonists; dopamine (DA), p-tyramine, (R)- and (S)-5-OH-dipropylaminotetralin (DPAT). We further probed the role of the conserved serine S1935.42 by mutagenesis, taking advantage of the preferential interaction of (S)-, but not (R)-5-OH-DPAT with this residue. Results suggested similar koffs for the two 5-OH-DPAT enantiomers at wild-type (WT) D2R, both being slower than the koffs of DA and p-tyramine. Conversely, the kon of (S)-5-OH-DPAT was estimated to be higher than that of (R)-5-OH-DPAT, in agreement with the higher potency of the (S)-enantiomer. Furthermore, S1935.42A mutation lowered the kon of (S)-5-OH-DPAT and reduced the potency difference between the two 5-OH-DPAT enantiomers. Kinetic Kds derived from the koff and kon estimates correlated well with EC50 values for all four compounds across four orders of magnitude, strengthening the notion that our assay captured meaningful information about binding kinetics. The approach presented here may thus prove valuable for characterizing D2R agonist candidate drugs.