Table 1.

Functional profiling of 5-HT_2B receptor mutants

Mutant	Agonist	Assay	WT EC50 (pEC50 ± SE)	Mutant EC50 (pEC50 ± SE)	Δlog(Emax/EC50)
D135L	5-HT	Ca2+	0.167 nM	no activity	n/a
			(9.78 ± 0.08)
	(R)-PZQ	Ca2+	2,000 nM	5,200 nM	−0.47
			(5.70 ± 0.05)	(5.28 ± 0.11)
	5-HT	BRET	0.97 nM	no activity	n/a
			(9.01 ± 0.22)
	(R)-PZQ	BRET	12,100 nM	14,300 nM	−0.24
			(4.92 ± 0.21)	(4.84 ± 0.20)
D135N	5-HT	Ca2+	0.167 nM	754 nM	−3.68
			(9.78 ± 0.08)	(6.12 ± 0.04)
	(R)-PZQ	Ca2+	2,000 nM	11,000 nM	−0.83
			(5.70 ± 0.05)	(4.95 ± 0.04)
	5-HT	BRET	0.97 nM	13,800 nM	−4.17
			(9.01 ± 0.22)	(4.86 ± 0.10)
	(R)-PZQ	BRET	12,100 nM	26,500 nM	−0.88
			(4.92 ± 0.21)	(4.58 ± 0.57)
D135A	5-HT	Ca2+	0.167 nM	95,300 nM	−5.75
			(9.78 ± 0.08)	(4.02 ± 0.13)
	(R)-PZQ	Ca2+	2,000 nM	141 nM	1.12
			(5.70 ± 0.05)	(6.85 ± 0.10)
	5-HT	BRET	0.97 nM	no activity	n/a
			(9.01 ± 0.22)
	(R)-PZQ	BRET	12,100 nM	374 nM	1.32
			(4.92 ± 0.21)	(6.43 ± 0.22)
S139A	5-HT	Ca2+	0.278 nM	0.978 nM	−0.54
			(9.56 ± 0.06)	(9.01 ± 0.04)
	(R)-PZQ	Ca2+	1,720 nM	247 nM	0.91
			(5.77 ± 0.08)	(6.61 ± 0.03)
	5-HT	BRET	0.97 nM	35.0 nM	−1.32
			(9.01 ± 0.22)	(7.46 ± 0.13)
	(R)-PZQ	BRET	15,400 nM	13,900 nM	0.36
			(4.81 ± 0.23)	(4.86 ± 0.15)
S139C	5-HT	Ca2+	0.278 nM	1.85 nM	−0.89
			(9.56 ± 0.06)	(8.70 ± 0.18)
	(R)-PZQ	Ca2+	1,720 nM	1,030 nM	0.28
			(5.77 ± 0.08)	(5.99 ± 0.06)
	5-HT	BRET	0.97 nM	10.6 nM	−1.05
			(9.01 ± 0.22)	(7.97 ± 0.19)
	(R)-PZQ	BRET	15,400 nM	11,100 nM	0.32
			(4.81 ± 0.23)	(4.95 ± 0.19)
S139V	5-HT	Ca2+	0.278 nM	38.9 nM	−1.46
			(9.56 ± 0.06)	(7.40 ± 0.56)
	(R)-PZQ	Ca2+	1,720 nM	1,530 nM	0.13
			(5.77 ± 0.08)	(5.82 ± 0.12)
	5-HT	BRET	0.97 nM	no activity	n/a
			(9.01 ± 0.22)
	(R)-PZQ	BRET	15,400 nM	6,600 nM	0.59
			(4.81 ± 0.23)	(5.18 ± 0.28)
F340L	5-HT	Ca2+	0.77 nM	1,087 nM	−3.43
			(9.11 ± 0.14)	(5.96 ± 0.02)
	(R)-PZQ	Ca2+	3,310 nm	no activity	n/a
			(5.48 ± 0.11)
F341L	5-HT	Ca2+	0.77 nM	no activity	n/a
			(9.11 ± 0.14)
	(R)-PZQ	Ca2+	3,310 nM	no activity	n/a
			(5.48 ± 0.11)
L209A	5-HT	Ca2+	0.278 nM	0.441 nM	−0.24
			(9.56 ± 0.06)	(9.36 ± 0.08)
	(R)-PZQ	Ca2+	1,720 nM	>150 µM	< −2.00
			(5.77 ± 0.08)	(<4.00)
	5-HT	BRET	0.972 nM	2.09 nM	−0.34
			(9.01 ± 0.22)	(8.68 ± 0.18)
	(R)-PZQ	BRET	15,400 nM	no activity	n/a
			(4.81 ± 0.23)
N344A	5-HT	Ca2+	0.278 nM	14.2 nM	−0.80
			(9.56 ± 0.06)	(7.85 ± 0.09)
	(R)-PZQ	Ca2+	1,720 nM	2,480 nM	−0.29
			(5.77 ± 0.08)	(5.61 ± 0.07)
	5-HT	BRET	0.972 nM	378 nM	−2.56
			(9.01 ± 0.22)	(6.42 ± 0.14)
	(R)-PZQ	BRET	15,400 nM	11,100 nM	0.13
			(4.81 ± 0.23)	(4.95 ± 0.14)

EC₅₀ values reported as mean (pEC₅₀ ± SE) for the wild-type and mutant 5-HT_2B receptor in response to 5-hydroxytryptamine (5-HT) and (R)-praziquantel [(R)-PZQ] are shown. Δlog(E_max/EC₅₀) (where E_max is maximum effect) calculated as log(E_max/EC₅₀)_mutant − log(E_max/EC₅₀)_WT. Data represent n ≥ 3 independent experiments for all assays. BRET, bioluminescence resonance energy transfer; n/a, not available.