Table S1. Content List of Extraction Buffers for Gpcrs

Table S1. Content list of extraction buffers for GPCRs.

Receptor / Detergent / Salt / pH buffer / Ligand / Additives
rhodopsin1F88 / 0.5-0.65% HPTO,
NG/ protein ratio ~2.2(w/w) / 90-120mM
Zn-acetate / 30-50 mM MES, pH 6.3-6.4
Na-acetate, pH 6.0 / N/A / N/A
β2AR2RH1 / 1% DDM / 500mM NaCl / 20 mM Tris, pH7.4 / N/A / 10 mg/ml benzamidine,
10 mg/ml leupeptin,
200 μmol PMSF
native opsin3CAP / 1% OG / N/A / N/A / N/A / N/A
β1AR2VT4 / 0.5% DDM / 350mM NaCl / 20 mM Tris-HCl, pH8 / N/A / 1 mM EDTA
A2AAR3EML / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 10 mM MgCl2
20 mM KCl / 10 mM HEPES, pH 7.5 / 4 mM theophylline / 2 mg/mL iodoacetamide, PIC
D3R3PBL / 1% (w/v) DDM, 0.2% (w/v) CHS / 10 mM MgCl2
20 mM KCl / 10 mM HEPES, pH 7.5 / 200 μM eticlopride / 2 mg/ml iodoacetamide, PIC
CXCR43ODU / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 500 mM NaCl / 50 mM HEPES, pH 7.5 / 100 μM IT1t / PIC
β1AR2Y00 / 0.5% DDM / 350mM NaCl / 20 mM Tris-HCl, pH8 / N/A / 1 mM EDTA
A2AAR3QAK / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 10 mM MgCl2
20 mM KCl / 10 mM HEPES, pH 7.5 / N/A / 2 mg/ml iodoacetamide,PIC
H1R3RZE / 1% (w/v) DDM, 0.2% (w/v) CHS / 500 mM NaCl / 20 mM HEPES, pH 7.5 / 5 mM doxepin / 20% (v/v) glycerol,
10 mg/ml iodoacetamide, PIC
β2AR3SN6 / 1% DDM / 500 mM NaCl / 20 mM Tris, pH 7.4 / N/A / 10 mg/ml benzamidine,
10 mg/ml leupeptin,
200 μM PMSF
S1P1R3V2Y / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 10 mM MgCl2
20 mM KCl / 10 mM HEPES, pH 7.5 / 250 μM ML056 / 2 mg/ml iodoacetamide,
40% glycerol, PIC
M3R4DAJ / 1% DDM, 0.03% CHS / 750mM NaCl / 30 mM HEPES, pH 7.5 / 1 μM titropium bromide / 30% glycerol,
2 mg/ml iodoacetamide
M2R3UON / digitonin/Na-cholate / 50 mM NaCl / 10 mM KPB, pH 7.0 / N/A / 1 mM EDTA, PIC
κ-OR4DJH / 1% (w/v) DDM, 0.2% (w/v) CHS / 150 mM NaCl / 50 mM HEPES, pH 7.5 / 20 μM naltrexone / 2 mg/ml iodoacetamide, PIC
μ-OR4DKL / 0.5% DDM, 0.3% CHAPS, 0.03% CHS / 500mM NaCl / 20 mM HEPES, pH7.5 / 1 μM naloxone / 30% (v/v) glycerol,
2 mg/ml iodoacetamide,
100 μM TCEP
NOP4EA3 / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 500mM NaCl
20 mM KCl / 50 mM HEPES, pH 7.5 / 25μM C-24 / 5% (v/v) glycerol,
1 mg/ml iodoacetamide
δ-OR4EJ4 / 1.0% LMNG, 0.3% Na-cholate, 0.03% CHS / 500mM NaCl / 20 mM HEPES, pH 7.5 / 1 μM naltrindole / 30% (v/v) glycerol,
2 mg/ml iodoacetamide
NTSR14GRV / 1% LMNG/0.1% CHS / 500 mM NaCl / 50 mM Tris-HCl, pH 7.4 / 10 μM NTS / 30% glycerol
PAR13VW7 / 1% DDM, 0.03% CHS,
0.2% Na-cholate / 350 mM NaCl / 20 mM Tris-HCl, pH7.5 / 100 nM vorapaxar / 10% glycerol,
2 mg/ml iodoacetamide
5-HT1B4IAR / 1% (w/v) DDM, 0.2% (w/v) CHS / 200 mM NaCl / 50 mM HEPES, pH 7.5 / 50 μM ERG / 2 mg/ml iodoacetamide, PIC
SMO4JKV / 1% (w/v) DDM, 0.2% (w/v) CHS / 200 mM NaCl / 50 mM HEPES, pH 7.5 / 30μM LY2940680 / 2 mg/ml iodoacetamide, PIC
CRF1R4K5Y / 2% (w/v) DM / 500 mM NaCL / 50 mM Tris-HCl, pH 8.0 / 0.3μM CP-376395 / N/A
GCGR4L6R / 1% (w/v) DDM, 0.1% (w/v) CHS / 166 mM NaCl / 25 mM HEPES, pH 7.0 / 270 μM NNC0640 / 13.3% glycerol, PIC
CCR54MBS / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 150 mM NaCl / 50 mM HEPES, pH7.5 / 100 μM Maraviroc / 2 mg/ml iodoacetamide, PIC
5-HT2B4IB4 / 1% (w/v) DDM, 0.2% (w/v) CHS / 150 mM NaCl / 10 mM HEPES, pH 7.5 / 50 μM ERG / PIC
β1AR4GPO / 2% DDM / 350mM NaCl / 20 mM Tris-HCl, pH 8 / N/A / PIC
M2R4MQS / 1% DDM, 0.03% CHS / 750mM NaCl / 30 mM HEPES, pH 7.5 / 1μM atropine / 30% glycerol,
2 mg/ml iodoacetamide
δ-OR4N6H / 0.75% (w/v) DDM, 0.15% (w/v) CHS / 500 mM NaCl / 50 mM HEPES, pH 7.5 / 25 μM naltrindole / N/A
mGluR14OR2 / 1% (w/v) DDM, 0.2% (w/v) CHS / 200 mM NaCl / 50 mM HEPES, pH 7.5 / 15 μM FITM / 2 mg/ml iodoacetamide, PIC
P2Y12R4NTJ / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 1 M NaCl
10 mM MgCl2
20 mM KCl / 50 mM HEPES, pH 7.5 / 200 μM AZD1283 / 2 mg/ml iodoacetamide, PIC
P2Y12R4PXZ / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 1 M NaCl
10 mM MgCl2
20 mM KCl / 50 mM HEPES, pH 7.5 / 20 μM 2MeSADP or 2MeSATP / 2 mg/ml iodoacetamide, PIC
mGluR54OO9 / 1.5% (w/v) DDM / 250 mM NaCl / 40 mM HEPES, pH 7.5 / 40 μM mavoglurant / 8 mM iodoacetamide, PIC
GPR404PHU / 1% (w/v) DDM, 0.1% (w/v) CHS / 500 mM NaCl / 20 mM HEPES, pH 7.6 / 1 μM TAK-875 / 10 mM imidazole, PIC,
1 mg/ml iodoacetamide
SMO4QIN / 1% (w/v) DDM, 0.2% (w/v) CHS / 200 mM NaCl / 50 mM HEPES, pH 7.5 / 15 μM SAG1.5 / PIC, 1 mg/ml iodoacetamide
US284XT1 / 1% (w/v) DDM, 0.2% (w/v) CHS / 150 mM NaCl / 10 mM HEPES, pH 7.4 / N/A / 20% (v/v) glycerol,
2 mg/ml iodoacetamide
OX2R4S0V / 1% (w/v) DDM, 0.2% Na-cholate,
0.2% CHS / 500 mM NaCl / 50 mM Tris, pH7.5 / 5 μM suvorexant / 10% glycerol,
2 mg/ml iodoacetamide
P2Y1R4XNV / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 300 mM NaCl / 50 mM HEPES, pH7.5 / N/A / 500 μM ATP, PIC, 2 mg/ml iodoacetamide
AT1R4YAY / 1% (w/v) DDM, 0.2% (w/v) CHS / N/A / N/A / N/A / N/A
rhodopsin4ZWJ / 0.5% (w/v) DDM, 0.1% (w/v) CHS / 100 mM NaCl / 20 mM Tris-HCl, pH7.4 / N/A / 10% glycerol, PIC
μ-OR5C1M / 0.5% DDM, 0.3% CHAPS,
0.03% CHS / 500mM NaCl / 20 mM HEPES, pH7.5 / 1 μM naloxone / 30% (v/v) glycerol,
2 mg/ml iodoacetamide,
100 μM TCEP
OX1R4ZJ8 / 1% (w/v) DDM, 0.2% Na-cholate, 0.2% (w/v) CHS / 500mM NaCl / 50 mM Tris-HCl, pH 7.4 / 5 μM SB-674042 / 10% glycerol,
2 mg/ml iodoacetamide
M1R5CXV / 1% DDM, 0.03% CHS / 750mM NaCl / 30 mM HEPES, pH 7.5 / 1 μM titropium bromide
1 μM atropine / 30% glycerol
M4R5DSG / 1% DDM, 0.03% CHS / 750mM NaCl / 30 mM HEPES, pH 7.5 / 1 μM titropium bromide
1μM atropine / 30% glycerol

Table S2. List of crystallization conditions.

Receptor / Concentration (mg/ml) / Detergent
(w/v) / Method / Temperature
(oC) / Time / Drop
(nl) / Precipitant
(nl) / Size
(μm)
rhodopsin1F88 / 7-9 / 0.55-0.75% HPTO + 0.4-0.55% NG / HDVD / 3-5 / 3 months / N/A / N/A / 100×10×50
β2AR2RH1 / 50 / 0.1% DDM / LCP / 21-23 / N/A / 25 or 50 / 800 / 30×15×5
native opsin3CAP / 5.0 / 1% OG / HDVD / 4 / 7 days / 2,000 / 2,000 / 100×100×200
β1AR2VT4 / 6.0 / 0.02% DM / HDVD / 18 / N/A / equal volume / 240×40×10
A2AAR3EML / 70 / 0.05% DDM + 0.001% CHS / LCP / 20 / N/A / 50 / 800 / 100×10×5
D3R3PBL / 20-30 / 0.05% DDM + 0.01% CHS / LCP / 20 / N/A / 40-50 / 800 / 45×30×10
CXCR43ODU / 60-70 / 0.05% DDM + 0.01% CHS / LCP / 20 / 10 days / 40-50 / 800 / 90×40×5
β1AR2Y00 / 14.5 / 0.45-1.8 mg/ml CHS + 0.5-0.65% Hega-10 / SDVD / 4 / NA / 200 / N/A / N/A
A2AAR3QAK / 60 / 0.025% DDM + 0.005% CHS / LCP / 20 / 7 days / 40 / 800 / 150×20×5
H1R3RZE / 30-40 / 0.025% DDM+0.005% CHS / LCP / 20 / N/A / 40-50 / 800 / 25-30
β2AR3SN6 / 50 / 0.1% DDM (β2AR-agonist)
0.02% LMNG (β2AR-Gs) / LCP / 20 / 3-4 days / 40 / 800 / 250
S1P1R3V2Y / 100 / 0.05% DDM + 0.001% CHS / LCP / 14 (1 day)
17 (2 days) / 3 days / 25 / 800 / 50×10×5
M3R4DAJ / 60 / 0.01% LMNG + 0.001% CHS / LCP / N/A / 2-3 days / 15 / 600 / 20
M2R3UON / 50 / 1% LMNG / LCP / 20 / 3-4 days / 40 / 800 / N/A
κ-OR4DJH / 40 / 0.03% DDM + 0.006% CHS / LCP / 20 / N/A / 45 / 800 / 60×20×10
μ-OR4DKL / 30 / 0.01% LMNG + 0.001% CHS / LCP / N/A / 5 days / 80 / 700 / 20
NOP4EA3 / 40 / 0.025% DDM + 0.005% CHS / LCP / 20 / 14 days / 40 / 800 / 40×10×3
δ-OR4EJ4 / 50 / 0.01% LMNG + 0.001% CHS / LCP / N/A / 5 days / 20-55 / 700 / 10-20
NTSR14GRV / 20-25 / 0.003% LMNG + 0.0003%CHS / LCP / N/A / 7 days / 65 / 750 / N/A
PAR13VW7 / 40-50 / 0.1% DDM + 0.02% CHS / LCP / N/A / 7 days / 20-40 / 700 / 20
5-HT1B4IAR / 50-80 / 0.05% DDM + 0.01% CHS / LCP / 20 / 2-3 days / 40 / 800 / 20-50
SMO4JKV / 50-60 / 0.03% DDM + 0.006% CHS / LCP / 20 / 2-3 days / 40 / 800 / 100×20×20
CRF1R4K5Y / 20-30 / 0.15% DM / LCP / 22.5 / N/A / 40-60 / 750 / 20-30
GCGR4L6R / 80 / 0.02% DDM + 0.004% CHS / LCP / 20 / 5 days / N/A / N/A / 50–100
CCR54MBS / 40-50 / 0.05% DDM + 0.01% CHS / LCP / 20 / 14 days / 45 / 800 / 100×20×15
5-HT2B4IB4 / 50 / 0.05% DDM + 0.01% CHS / LCP / 20 / 3 days / 40 / 800 / 100×30×20
β1AR4GPO / 8 / 0.02% DDM / HDVD / 18 / N/A / 1,000 / 1,000 / N/A
M2R4MQS / N/A / 0.01% LMNG + 0.001% CHS / LCP / N/A / 2 days / 30-100 / 600 / N/A
δ-OR4N6H / 20 / 0.02% DDM + 0.004% CHS / LCP / 20 / 10 days / 40 / 800 / 50×10×3
mGluR14OR2 / 50-80 / 0.05 % DDM + 0.01% CHS / LCP / 20 / 5-6 days / N/A / N/A / 35×35×15
P2Y12R4NTJ / ~40 / 0.05% DDM + 0.01% CHS / LCP / N/A / 2 weeks / 40 / 800 / 150×15×15
P2Y12R4PXZ / 30 / 0.05% DDM + 0.01% CHS / LCP / N/A / 2 weeks / 40 / 800 / 80×50 ×5
mGluR54OO9 / ~25 / 0.03% DDM / LCP / 20 / N/A / 40-60 / 750 / 40
GPR404PHU / 25-35 / 0.02% LMNG + 0.0003% CHS / LCP / 20 / 1-2 days / 100 / 800 / 25
SMO4QIN / 50-60 / 0.03% DDM + 0.006% CHS / LCP / 20 / N/A / 50 / 1,000 / N/A
US284XT1 / 34 / 0.02% DDM + 0.004% CHS / LCP / 20 / 3-5 days / 30 / 700 / 20
OX2R4S0V / 30 / 0.05% LMNG + 0.02% CHS / LCP / 20 / 3 days / 40 / 800 / 20
P2Y1R4XNV / N/A / 0.05% DDM + 0.01% CHS / LCP / N/A / 2 weeks / 40-50 / 800 / 100-130
AT1R4YAY / 30 / N/A / LCP / 20 / 6.5 hours / SFX (65,000) / 10×2×2
rhodopsin4ZWJ / 40-50 / 0.005% LMNG + 0.001% CHS / LCP / 20 / 4 days / 50 / 800 / 5-10
μ-OR5C1M / 50 / 0.01% LMNG + 0.001% CHS / LCP / N/A / 7 days / 30 / 500 / N/A
OX1R4ZJ8 / >30 / 0.05% LMNG / LCP / 20 / 7 days / 40 / 800 / 40
M1R5CXV / 50 / 0.01% LMNG + 0.001% CHS / LCP / 20 / 24 hours / 50 / 800 / N/A
M4R5DSG / 50 / 0.01% LMNG + 0.001% CHS / LCP / 20 / 24 hours / 20-40 / 600 / N/A

Table S3. Content list of precipitants for crystallization.

Receptor / PEG (v/v) / Salt / pH buffer / Ligand / Additives
rhodopsin1F88 / N/A / 65-90 mM ZnOAc
840-860 mM (NH4)2SO4 / 30-50 mM MES, pH 6.3-6.4
NaOAc, pH 6.0 / N/A / 5-7 mMβ-mercaptoethanol,
0.55-0.75% HPTO,
0.45-0.55% NG
β2AR2RH1 / 30-35% PEG400 / 100-200 mM Na2SO4 / 100 mM Bis-Tris propane, pH 6.5-7.0 / N/A / 5-7% (v/v) 1,4-butanediol
native opsin3CAP / N/A / N/A / N/A / N/A / N/A
β1AR2VT4 / 29-32% PEG600 / N/A / 100 mM ADA:NaOH, pH 6.9-7.3 / N/A / N/A
A2AAR3EML / 30% PEG400 / 186 mM Li2SO4 / 100 mM Nacitrate, pH 6.5 / 200 μM ZM241385 / N/A
D3R3PBL / 26-34% PEG400 / 200-350 mM NH4OAc / 100 mM Bis-Tris propane, pH 7.5-8.0 / 1mM eticlopride / 2-3% (w/v) D-(+)-Glucose
CXCR43ODU / 20% PEG400 / 300 mM Namalonate / 100 mM Nacitrate, pH 5.5 / 2 mM IT1t / 5 mM Taurine
β1AR2Y00 / 25%PEG600 / N/A / 100mM Tris-HCl, pH 8.5
100mM Bicine, pH9.0 / N/A / N/A
A2AAR3QAK / 30% PEG400 / 150-200 mM MgCl2 / 100 mM Nacitrate, pH 5.0-5.5 / N/A / N/A
H1R3RZE / 26-30% PEG400 / 300 mM (NH4)3PO4,
10 mM MgCl2 / 100 mM Nacitrate, pH 4.5 / 1 mM doxepin / N/A
β2AR3SN6 / 18-22% PEG400 / 350-450 mM KNO3 / 100 mM MES, pH6.5 / 10 μM BI-167107 / 1 mM TCEP, 10 mM foscarnet
S1P1R3V2Y / 34-36% PEG400 / 80 mM Nacitrate / 100 mM Tricine, pH 7.75 / N/A / 4% (v/v) glycerol
M3R4DAJ / 27-38% PEG300 / 100 mM (NH4)3PO4 / 100 mM HEPES, pH 7.5 / N/A / 1% (w/w) 1,2,3-heptanetriol
M2R3UON / 25-35% PEG300 / 100 mM (NH4)3PO4 / 100 mM HEPES, pH 7.0-7.8 / N/A / 2% 2-methyl-2,4-pentanediol
κ-OR4DJH / 28-32% PEG400 / 350-450 mM KNO3 / 100 mM Nacitrate, pH 5.8-6.4 / N/A / N/A
μ-OR4DKL / 30-38% PEG400 / 300 mM Li2SO4 / 100 mM HEPES, pH7.0 / N/A / 7.5% DMSO
NOP4EA3 / 25-30% PEG400 / 100-200 mM KNatartrate / 100 mM Bis-Tris propane, pH6.4 / N/A / N/A
δ-OR4EJ4 / 29-33% PEG400 / 120-180 mM Nacitrate/
350 mM MgCl2 / 100 mM HEPES, pH 7.5 / N/A / N/A
NTSR14GRV / 20.8% PEG400 / 43 mM KNatartrate / 80 mM HEPES, pH 7.0 / N/A / 2 mM TCEP
PAR13VW7 / 25-35% PEG300 / 100-200 mM NaCl / 100 mM Na3PO4, pH 6.0-6.5 / N/A / N/A
5-HT1B4IAR / 30% PEG400 / 400 mM LiCl / 100 mM Tris-HCl, pH 7.5 / N/A / N/A
SMO4JKV / 32% PEG400 / 70 mM NH4F / 100mM HEPES, pH7.8 / N/A / 4-8% (v/v)
polypropylene glycol P 400
CRF1R4K5Y / 30% PEG400 / 200 mM Li2SO4 / 100 mM Nacitrate, pH 5.5 / 0.6 μM CP-376395 / N/A
GCGR4L6R / 9-17% PEG400 / 140-200 mM KNatartrate / 100 mM MES, pH 6.0 / 200 μM NNC0640 / 0.35-0.55% (v/v)
Jeffamine M-600
CCR54MBS / 32-38% PEG400 / 100-200 mM NaCl / 100 mM HEPES, pH 7.0 / 1 mM Maraviroc / N/A
5-HT2B4IB4 / 30% PEG400 / 50-75 mM MgSO4 / 100 mM Tris-HCl, pH 8.0-8.5 / N/A / N/A
β1AR4GPO / 26-30% PEG200 / 100-300 mM (NH4)2SO4 / 20 mM Nacitrate, pH 3.6-4 / N/A / N/A
M2R4MQS / 10-20% PEG300 / N/A / 100 mM HEPES, pH7.2-7.9 / N/A / 1.2% 1,2,3-heptanetriol,
20-80 mM EDTA
δ-OR4N6H / 31-34% PEG400 / 95-120 mM KNatartrate / 100 mM MES, pH 6.1-6.2 / 1 mM naltrindole / 5% (v/v) ethylene glycol
mGluR14OR2 / 27-30% PEG400 / 80-120 mM(NH4)3HPO4 / 100 mM HEPES, pH 7.0 / N/A / 4-8 mM TCEP
P2Y12R4NTJ / 35% PEG400 / 50-150 mM NH4HCO2 / 100 mM Nacacodylate, pH 6.0-6.5 / 200 μM AZD1283 / N/A
P2Y12R4PXZ / 30-40% PEG400 / 300-450 mM NH4OAc / 100 mM Nacitrate, pH5.0 / 500 μM 2MeSADP / 3% v/v 1-propanol
mGluR54OO9 / 24-34% PEG400 / 100-250 mM (NH4)3HPO4 / 100 mM MES, pH 5.5-6.8 / 50 μM mavoglurant / N/A
GPR404PHU / 29-31% PEG400/
39.8% PEG400 / 200 mM Namalonate/
100 mM (NH4)3PO4 / 100 mM Tris-HCl, pH 8.0/
100 mM Bis-Tris propane, pH 7.2 / N/A / N/A
SMO4QIN / 30% PEG400 / 100 mM MgSO4 / 100 mM MES, pH6.0 / N/A / 2-3% polypropylene glycol P 400
US284XT1 / 32% PEG300 / 100 mM Nasuccinate,
pH 7.0 / 100mM MES, pH 6.3 / N/A / 1% (v/v) polypropylene glycol P 400
OX2R4S0V / 31% PEG400 / 200 mM Naformate / 100 mM Nacitrate, pH 5.9 / N/A / 3% 2,5-hexanediol
P2Y1R4XNV / 0-10% PEG2000 MME / 100-300 mM (NH4)3HPO4 / 100 mM Nacitrate, pH 6.5 / 50 μM BPTU / N/A
AT1R4YAY / 28% PEG400 / 450 mM (NH4)3PO4 / 100 mM Nacitrate, pH5.0 / N/A / 4% (v/v) DMSO
rhodopsin4ZWJ / 28% PEG400 / 50 mM MgOAc / 50 mM Nacitrate, pH 5.0 / N/A / N/A
μ-OR5C1M / 15-25% PEG300 / N/A / 100 mM HEPES, pH 7.0-7.5 / N/A / 1% 1,2,3-heptanetriol,
0.5-1.0% polypropylene glycol P 400
OX1R4ZJ8 / 32% PEG400 / 200 mM HCO₂K / 100 mM Nacitrate, pH 5.6 / N/A / 4% 1,4-butanediol
M1R5CXV / 33% PEG300 / 100 mM NaOAc / 100 mM Bis-Tris propane, pH 8.0 / N/A / N/A
M4R5DSG / 25-40% PEG300 / N/A / 100 mM MES, pH 5.5-6.5 / N/A / 50-100 mM EDTA, pH 8.0