SUPPLEMENTAL INFORMATION Identification and Analysis of Hepatitis C virus NS3 Helicase

Inhibitors Using Binding Assays

Sourav Mukherjee,1 Alicia M. Hanson,1 William R. Shadrick,1 Jean Ndjomou,1 Noreena L.

Sweeney,1 John J. Hernandez,1 Diana Bartczak,1 Kelin Li,2 Kevin J. Frankowski,2 Julie A.

Heck,3,4 Leggy A. Arnold,1 Frank J. Schoenen,3 and David N. Frick1,*

1Department of Chemistry and Biochemistry, University of Wisconsin-Milwaukee,

Milwaukee, WI, 53211, USA. 2University of Kansas Specialized Chemistry Center,

University of Kansas, 2034 Becker Dr., Lawrence, KS 66047. 3Department of

Biochemistry and Molecular Biology, New York Medical College, Valhalla, NY, 10595.

Table S1: Compound effects on the fluorescent polarization (FP)-based NS3h-DNA binding assay and the molecular beacon based helicase assay (MBHAs). Means ± standard deviations (n=2) are reported for normalized inhibition (%) and assay interference (ratio). Compounds colored blue are primuline derivatives, green compounds are known DNA binding agents, and pink compounds are other disclosed NS3 helicase inhibitors. [Data relate to Fig. 4]

Table S2: Results of a screen of Sigma’s LOPAC for inhibitors of Cy5-dT15-NS3h complex formation. Pink compounds inhibited complex formation more than 60%, and green compounds interfered with the assay more than 20%. [Data relate to Fig. 5]

Table S3: The Cy-15-dT15-SSB binding assay as a screen for specific HCV helicase inhibitors. Means ± standard deviations of IC50 values obtained in titrations (n=3) with each of the shown primuline derivatives. Compounds are ranked based on their ability to inhibit the formation of the Cy-15-dT15-SSB complex, with the least potent compounds on the top. [Data relate to Fig. 8]! Table&S1

$Cy52dT152NS3h$Binding$ $Cy52dT152NS3h$Binding$ DNA$Unwinding$(MBHA)$ DNA$Unwinding$(MBHA)$ CID$No./Compound$Name$ %$Inhibition Interference$ %Inhibition Interference TO*PRO*3 *63&±&0 1.4&±&0.0 115&±&2 0.5&±&0.0 Hoechst&33258 *96&±&2 0.1&±&0.0 104&±&6 0.3&±&0.0 Chromomycin&A3 *4&±&8 1.0&±&0.0 96&±&1 1.0&±&0.0 4049406 62&±&5 0.7&±&0.0 96&±&2 0.7&±&0.0 53308659 17&±&2 1.1&±&0.1 95&±&0 1.0&±&0.0 SYBR&Green&I 24&±&2 0.8&±&0.0 91&±&3 0.7&±&0.0 53255474 46&±&65 0.3&±&0.2 90&±&2 0.4&±&0.0 44251438 74&±&17 1.0&±&0.0 89&±&3 0.9&±&0.0 486270 30&±&4 0.6&±&0.0 87&±&4 0.6&±&0.0 53239937 9&±&2 1.0&±&0.0 87&±&5 1.0&±&0.0 44251434 47&±&10 1.0&±&0.0 82&±&4 1.0&±&0.0 44251428 54&±&4 1.0&±&0.0 81&±&12 1.0&±&0.0 247520 *17&±&2 0.8&±&0.0 80&±&4 0.7&±&0.0 3413238 63&±&2 0.7&±&0.0 78&±&7 0.7&±&0.0 49849282 22&±&3 1.0&±&0.0 77&±&8 1.0&±&0.0 Titan&Yellow 97&±&4 1.1&±&0.0 75&±&3 1.0&±&0.0 483665 *21&±&2 0.9&±&0.0 74&±&4 0.8&±&0.0 4176595 *13&±&3 0.8&±&0.0 69&±&2 0.9&±&0.0 4111820 4&±&2 0.7&±&0.0 68&±&7 0.9&±&0.0 7080057 *17&±&0 0.9&±&0.0 67&±&1 0.9&±&0.0 49849300 73&±&0 1.0&±&0.0 63&±&2 1.0&±&0.0 Soluble&Blue&HT 78&±&0 1.0&±&0.0 63&±&12 0.8&±&0.0 52939982 *9&±&2 0.9&±&0.0 59&±&4 1.0&±&0.0 44251431 73&±&8 1.0&±&0.0 58&±&23 1.0&±&0.0 Ellipticine *6&±&0 1.0&±&0.0 57&±&4 0.6&±&0.0 49849299 *2&±&1 1.0&±&0.0 53&±&14 1.0&±&0.0 Netropsin 8&±&1 0.9&±&0.0 51&±&20 0.8&±&0.1 Thiazole&Orange *4&±&1 0.9&±&0.0 49&±&7 0.7&±&0.0 50930741 56&±&13 1.0&±&0.0 46&±&5 1.0&±&0.0 50930749 *1&±&5 1.0&±&0.0 41&±&13 1.0&±&0.0 50930730 46&±&30 1.0&±&0.0 41&±&25 1.0&±&0.0 49849293 39&±&2 1.0&±&0.0 32&±&12 0.9&±&0.0 49849293 20&±&6 1.0&±&0.0 31&±&4 0.9&±&0.0 50930737 58&±&14 1.0&±&0.0 31&±&11 1.0&±&0.0 50930733 22&±&6 1.0&±&0.0 29&±&4 1.0&±&0.0 50930738 91&±&1 1.1&±&0.0 28&±&2 1.0&±&0.0 DAPI 4&±&0 0.9&±&0.0 28&±&9 0.8&±&0.0 Rebeccamycin *10&±&2 1.0&±&0.0 27&±&11 0.8&±&0.1 46913723 *10&±&4 1.0&±&0.0 27&±&3 0.9&±&0.0 Proflavin *4&±&1 0.9&±&0.0 27&±&4 0.8&±&0.0 53308658 30&±&14 0.9&±&0.2 27&±&2 1.0&±&0.0 50930755 18&±&3 1.0&±&0.0 26&±&1 0.9&±&0.0 49849287 19&±&5 1.0&±&0.0 25&±&1 0.9&±&0.0 Quinacrine *5&±&4 0.9&±&0.0 25&±&2 0.9&±&0.0 49849302 28&±&1 1.0&±&0.0 24&±&6 0.9&±&0.0 16060752 11&±&5 0.9&±&0.0 24&±&3 0.8&±&0.0 5479184 10&±&3 0.9&±&0.0 24&±&1 0.9&±&0.0 50930751 19&±&6 1.0&±&0.0 23&±&0 1.0&±&0.0 53312458 43&±&6 1.0&±&0.0 23&±&2 0.9&±&0.0 49849298 13&±&1 1.0&±&0.0 22&±&3 0.9&±&0.0 50930740 34&±&6 1.0&±&0.0 22&±&1 1.0&±&0.0 Thioflavin&T *8&±&0 1.0&±&0.0 21&±&2 0.9&±&0.0 3084034 30&±&4 1.0&±&0.0 21&±&12 0.8&±&0.0 Eva&Green *3&±&4 1.0&±&0.0 20&±&2 0.9&±&0.0 53255447 18&±&7 1.0&±&0.0 20&±&0 0.9&±&0.1 52914816 10&±&2 1.0&±&0.0 20&±&9 1.0&±&0.0 50930745 11&±&6 1.0&±&0.0 19&±&6 0.9&±&0.0 Berenil *46&±&0 1.0&±&0.0 19&±&3 0.9&±&0.0 49849280 41&±&0 1.0&±&0.0 17&±&3 0.9&±&0.0 46202556 *8&±&0 1.0&±&0.0 17&±&1 1.0&±&0.0 50930743 38&±&2 1.0&±&0.0 17&±&5 0.9&±&0.0 49849276 28&±&11 1.0&±&0.0 16&±&5 1.0&±&0.0 50930732 4&±&0 1.0&±&0.0 16&±&8 1.0&±&0.0 50930756 52&±&10 1.0&±&0.0 15&±&2 1.0&±&0.0 9585555 *4&±&2 0.9&±&0.0 15&±&5 1.0&±&0.0 3832647 *6&±&2 1.0&±&0.0 15&±&2 1.0&±&0.0 Trioxsalen *6&±&2 1.0&±&0.0 15&±&6 0.9&±&0.0 Carrier&Blank/DMSO *7&±&2 1.0&±&0.0 8&±&4 1.0&±&0.0 50930734 18&±&4 1.1&±&0.0 14&±&6 1.0&±&0.0 49849284 26&±&8 1.0&±&0.0 14&±&7 1.0&±&0.0

1 Table&S1

$Cy52dT152NS3h$Binding$ $Cy52dT152NS3h$Binding$ DNA$Unwinding$(MBHA)$ DNA$Unwinding$(MBHA)$ CID$No./Compound$Name$ %$Inhibition Interference$ %Inhibition Interference 53255452 13&±&5 1.0&±&0.0 14&±&0 0.9&±&0.0 5362502 *1&±&5 1.0&±&0.0 13&±&3 0.9&±&0.0 Luciferin *4&±&1 1.0&±&0.0 13&±&2 0.9&±&0.0 9682118 *9&±&1 1.0&±&0.0 13&±&4 1.0&±&0.0 3643004 *5&±&1 1.0&±&0.0 12&±&5 1.0&±&0.0 Carrier&Blank/DMSO *8&±&1 1.0&±&0.0 12&±&4 1.0&±&0.0 46916208 *9&±&2 1.0&±&0.0 11&±&4 1.0&±&0.0 5691857 3&±&0 0.9&±&0.0 11&±&4 1.0&±&0.0 4594423 *5&±&1 1.0&±&0.0 11&±&4 1.0&±&0.0 3496625 *5&±&0 1.0&±&0.0 11&±&3 1.0&±&0.0 2998553 *1&±&2 1.0&±&0.0 11&±&9 0.9&±&0.0 6540194 *9&±&0 1.0&±&0.0 10&±&4 1.0&±&0.0 Primulin 31&±&3 1.0&±&0.0 10&±&4 1.0&±&0.0 978069 1&±&1 0.9&±&0.0 10&±&8 1.0&±&0.0 5447034 *3&±&2 1.0&±&0.0 10&±&14 0.9&±&0.1 45382099 *4&±&1 1.0&±&0.0 10&±&2 1.0&±&0.0 6064372 *6&±&3 0.9&±&0.0 10&±&2 1.0&±&0.0 53255449 0&±&1 1.0&±&0.0 10&±&6 1.0&±&0.0 49849286 27&±&5 1.0&±&0.0 10&±&3 1.0&±&0.0 1293387 *7&±&1 0.9&±&0.0 10&±&7 1.0&±&0.0 6540199 *7&±&3 1.0&±&0.0 9&±&5 1.0&±&0.0 7314627 1&±&1 1.0&±&0.0 9&±&6 1.0&±&0.0 5281571 20&±&2 1.0&±&0.0 9&±&5 1.0&±&0.0 45382098 *4&±&3 1.0&±&0.0 9&±&3 1.0&±&0.0 49795072 *8&±&0 1.0&±&0.0 9&±&9 1.0&±&0.0 4473963 *11&±&1 1.0&±&0.0 8&±&2 1.0&±&0.0 44251428 24&±&6 1.0&±&0.0 8&±&2 1.0&±&0.0 16060753 *10&±&4 1.0&±&0.0 8&±&3 1.0&±&0.0 Carrier&Blank/DMSO *8&±&0 1.0&±&0.0 14&±&6 0.9&±&0.0 46916209 *9&±&0 1.0&±&0.0 8&±&5 1.0&±&0.0 49849289 36&±&2 1.0&±&0.0 8&±&6 1.0&±&0.0 5860596 *4&±&0 1.0&±&0.0 8&±&6 1.0&±&0.0 4611776 *5&±&3 1.0&±&0.0 8&±&2 1.0&±&0.0 16060655 *8&±&2 1.0&±&0.0 7&±&4 1.0&±&0.0 50930748 31&±&2 1.0&±&0.0 7&±&1 1.0&±&0.0 46839370 29&±&5 1.0&±&0.0 7&±&3 1.0&±&0.0 5736971 *3&±&2 0.9&±&0.0 7&±&2 1.1&±&0.0 1601813 *5&±&3 1.0&±&0.0 7&±&3 1.0&±&0.0 2186405 *6&±&0 0.9&±&0.0 7&±&11 1.0&±&0.0 6540163 *10&±&1 0.9&±&0.0 7&±&2 1.0&±&0.0 5293297 *7&±&2 0.9&±&0.0 7&±&0 1.0&±&0.0 45382102 *4&±&2 1.0&±&0.0 7&±&3 1.0&±&0.0 50904396 *10&±&1 1.0&±&0.0 6&±&13 1.0&±&0.0 2571817 *8&±&2 1.0&±&0.0 6&±&4 1.0&±&0.0 4371917 *26&±&4 1.0&±&0.0 6&±&6 1.0&±&0.0 2330223 *47&±&0 0.9&±&0.0 6&±&5 1.0&±&0.0 46897855 37&±&2 1.0&±&0.0 5&±&2 1.0&±&0.0 49795071 2&±&2 1.0&±&0.0 5&±&11 1.0&±&0.0 16238633 *4&±&1 1.0&±&0.0 5&±&4 1.0&±&0.0 1959044 *1&±&2 0.9&±&0.0 4&±&1 1.0&±&0.0 4051948 *7&±&1 1.0&±&0.0 4&±&11 1.0&±&0.0 2214934 *4&±&3 0.9&±&0.0 4&±&3 1.0&±&0.0 4872167 *4&±&2 1.0&±&0.0 4&±&5 1.0&±&0.0 49849294 32&±&6 1.0&±&0.0 4&±&5 1.0&±&0.0 45382104 0&±&1 1.0&±&0.0 4&±&3 1.0&±&0.0 49849279 *5&±&6 1.0&±&0.0 4&±&9 1.0&±&0.0 1126404 *8&±&2 1.0&±&0.0 3&±&6 1.0&±&0.0 53255450 12&±&2 1.0&±&0.0 3&±&4 1.0&±&0.0 49849295 8&±&3 1.0&±&0.0 3&±&1 1.0&±&0.0 45382101 *5&±&2 1.0&±&0.0 2&±&2 1.0&±&0.0 53255448 9&±&1 1.0&±&0.0 2&±&6 1.0&±&0.0 24818137 *3&±&2 1.0&±&0.0 1&±&7 1.0&±&0.0 44251427 8&±&2 1.0&±&0.0 0&±&16 1.1&±&0.0 44251433 7&±&3 1.0&±&0.0 0&±&11 1.1&±&0.0 2115891 7&±&2 0.9&±&0.0 *1&±&6 1.1&±&0.0 45382097 *7&±&2 1.0&±&0.0 *1&±&9 1.0&±&0.0 44251429 18&±&0 1.0&±&0.0 *1&±&3 1.1&±&0.0 46839371 *8&±&2 0.9&±&0.0 *1&±&8 1.0&±&0.0 49795073 *9&±&1 1.0&±&0.0 *1&±&0 1.0&±&0.0 45382103 0&±&2 1.0&±&0.0 *2&±&5 1.1&±&0.0

2 Table&S1

$Cy52dT152NS3h$Binding$ $Cy52dT152NS3h$Binding$ DNA$Unwinding$(MBHA)$ DNA$Unwinding$(MBHA)$ CID$No./Compound$Name$ %$Inhibition Interference$ %Inhibition Interference 3923766 *9&±&2 0.9&±&0.0 *2&±&8 1.0&±&0.0 3291242 *1&±&2 0.9&±&0.0 *4&±&14 1.1&±&0.0 3138378 *9&±&0 0.9&±&0.0 *5&±&4 1.0&±&0.0 49849290 1&±&0 1.0&±&0.0 22&±&7 1.0&±&0.0

3 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( Protoporphyrin IX disodium P 8293 140 2.0 Mitoxantrone dihydrochloride M 6545 119 0.5 Tyrphostin AG 538 T 7822 100 1.0 sodium salt S 2671 96 1.0 Aurintricarboxylic acid A 1895 96 1.0 NF 023 N 8652 91 1.1 Reactive Blue 2 R-115 86 0.7 PPNDS tetrasodium P 2738 58 0.8 MRS 2159 M 7684 43 1.0 2-Methylthioadenosine triphosphate tetrasodium A-023 41 1.0 I-OMe-Tyrphostin AG 538 T 7697 37 0.9 Tyrphostin AG 537 T 4693 37 1.0 2-Chloroadenosine triphosphate tetrasodium C-145 37 1.0 Aurothioglucose A0606 36 1.0 2-(Methylthio) 5'-diphosphate trisodium salt hydrate M152 33 0.9 Me-3,4-dephostatin M 9440 27 0.9 6-Hydroxy-DL-DOPA H 2380 22 0.9 Tyrphostin 51 T 7665 19 0.8 Tetraethylammonium chloride T 2265 14 1.0 JFD00244 J4829 13 1.1 L-745,870 hydrochloride L-131 12 1.0 Methoctramine tetrahydrochloride M-105 11 1.0 SCH-202676 hydrobromide S 4063 8 1.0 M 6760 8 1.0 SU 5416 S 8442 6 1.0 Cefotaxime sodium C 7912 6 1.0 L-750,667 trihydrochloride L-133 6 0.9 Ro 41-0960 R-108 5 0.9 Tyrphostin AG 879 T 2067 5 1.0 U-73122 U 6756 5 0.8 SKF 89626 S 3066 5 1.0 3-Aminopropionitrile fumarate A 3134 4 1.0 Cefsulodin sodium salt hydrate C 8145 4 1.0 DL-erythro-Dihydrosphingosine D 6908 4 0.9 Pentolinium di[L(+)-tartrate] P 3520 4 1.0 Lercanidipine hydrochloride hemihydrate L 6668 3 1.0 E9531 3 1.0 GYKI 52466 hydrochloride G-119 3 1.0 P1,P4-Di(adenosine-5')tetraphosphate triammonium D 1262 3 1.0 Hemicholinium-3 H-108 3 1.0 GW7647 G 6793 3 1.0 R(-)-Isoproterenol (+)-bitartrate I 2760 3 1.0 hydrochloride N 7510 3 1.0 Pyrilamine maleate P 5514 3 1.0 MK-912 M 7065 3 1.0 Tyrphostin 47 T 7540 3 0.9 IPA-3 I 2285 3 1.0 BIX 01294 trihydrochloride hydrate B9311 3 1.0 (+)- methanesulfonate B 2134 2 0.9 L- sulfate C 9758 2 1.0 DPO-1 D7443 2 1.0 (2S,1'S,2'S)-2-(carboxycyclopropyl) C-237 2 1.0 Decamethonium dibromide D 1260 2 1.0 N-Acetyl-2,3-dehydro-2-deoxyneuraminic acid D 9050 2 1.0 (±)-8-Hydroxy-DPAT hydrobromide H 8250 2 1.0 Dequalinium chloride hydrate D 3768 2 0.9 1,10-Diaminodecane D14204 2 1.0 Etoposide E 1383 2 1.1 7-Cyclopentyl-5-(4-phenoxy)phenyl-7H-pyrrolo[2,3-d]pyrimidin-4-ylamine C 8863 2 1.0 Emetine dihydrochloride hydrate E 2375 2 1.0 hemifumarate salt B 2185 2 1.0 BIO B 1686 2 0.9 Nimustine hydrochloride N 8659 2 1.0 (-)-Perillic acid 218359 2 0.9 (±)-threo-1-Phenyl-2-decanoylamino-3-morpholino-1-propanol hydrochloride P 7340 2 1.0 tartrate S-006 2 1.0 Rutaecarpine R 3277 2 1.0 L- T 0254 2 1.0

1 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( hydrochloride monohydrate Z2777 2 1.0 Tyrphostin AG 698 T 5193 2 0.9 Vincristine sulfate V 8879 2 1.0 dihydrate Q 0125 2 0.9 Succinylcholine chloride S 8251 2 1.0 xinafoate S 5068 2 1.0 DL-Buthionine-[S,R]-sulfoximine B 2640 1 1.0 hydrochloride A 7410 1 0.9 DL-p-Chlorophenylalanine methyl ester hydrochloride C 3635 1 1.0 63Methoxy31,2,3,43tetrahydro39Hpyrido[3,4b]& 291552 1 0.9 6-Aminohexanoic acid A 7824 1 0.9 CNS-1102 C 4238 1 1.0 hydrochloride B 7283 1 1.0 C 5270 1 1.0 6-Fluoronorepinephrine hydrochloride B-012 1 0.9 Calcimycin C 7522 1 1.0 sulfate A 7127 1 1.0 C 2755 1 1.0 SB 222200 S 5192 1 1.0 CK2 Inhibitor 2 C 7367 1 1.0 PD 169316 P 9248 1 1.0 (±)-CPP C-104 1 1.0 JX401 J4774 1 1.0 4-Hydroxy-3-methoxyphenylacetic acid H 1252 1 1.0 (-)- fumarate E-100 1 1.0 N,N-Dihexyl-2-(4-fluorophenyl)indole-3-acetamide D 8555 1 1.0 SP600125 S 5567 1 1.0 L-3,4-Dihydroxyphenylalanine D 9628 1 1.0 Chloro-IB-MECA C-277 1 1.0 D 1064 1 1.0 R(-)-2,10,11-Trihydroxy-N-propylnoraporphine hydrobromide D-030 1 1.0 AC-93253 iodide A9605 1 0.9 3,5-Dinitrocatechol D-131 1 1.0 hydrochloride H 9876 1 1.0 5,5-Diphenylhydantoin D 4007 1 1.0 4-Hydroxybenzhydrazide H 9882 1 1.1 Glybenclamide G 0639 1 1.0 hydrochloride G 1043 1 1.0 2,4-Diamino-6-pyrimidinone D1920-6 1 1.1 SCH-28080 S 4443 1 1.0 Diacylglycerol Kinase Inhibitor II D 5794 1 1.0 Metolazone M 1195 1 1.0 IMID-4F hydrochloride I 2279 1 1.0 MRS 1845 M 1692 1 0.9 Kenpaullone K 3888 1 1.0 NCS-356 N 4034 1 1.0 Pancuronium P 1918 1 1.0 (±)- (+)-tartrate M 5391 1 1.0 Cisplatin P 4394 1 0.8 LFM-A13 L 8789 1 1.0 NNC 55-0396 N 0287 1 1.0 NO-711 hydrochloride N-142 1 1.0 MDL 28170 M 6690 1 1.1 Pilocarpine nitrate P 6628 1 1.0 SIB 1757 S 9186 1 0.9 Pyrazinecarboxamide P 7136 1 1.0 Primidone P 7295 1 1.0 13-cis-retinoic acid R 3255 1 0.9 PAC-1 P0115 1 0.9 Trimethoprim T 7883 1 1.0 REV 5901 R 5523 1 0.9 Tranilast T 0318 1 1.0 Sobuzoxane S 4692 1 1.0 hydrochloride T 2057 1 0.9 P 9689 1 1.0 Sulfaphenazole S 0758 1 1.0 sulfate sodium P-162 1 1.0 hydrochloride X 6000 1 1.0

2 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( Quinacrine dihydrochloride Q 3251 1 1.0 hydrochloride T 6764 1 1.0 Sulindac S 8139 1 1.0 IMS2186 I 7160 1 1.0 Zonisamide sodium Z 2001 1 1.0 Zimelidine dihydrochloride Z-101 1 1.0 N3Phenylanthranilic&acid 144509 0 0.9 (±)3Nipecotic&acid 211672 0 0.9 N-Acetyl-L- A 7250 0 0.9 Cyclosporin A C 3662 0 0.9 Captopril C 4042 0 1.0 Acetamide A&0500 0 0.9 L- A 9256 0 0.9 C 4522 0 1.0 Carbetapentane citrate C 4662 0 1.0 Fulvestrant I4409 0 0.9 2-Chloroadenosine C 5134 0 1.0 GABA A&2129 0 1.0 (+)- hydrochloride D-033 0 1.0 8-(p-Sulfophenyl) A-013 0 1.0 Indirubin-3'- I0404 0 1.0 Cefaclor C 6895 0 1.0 C 0625 0 1.0 CGP-13501 C 0987 0 1.0 1-(4-Chlorobenzyl)-5-methoxy-2-methylindole-3-acetic acid C 1610 0 1.0 S-(+)-PD 123177 trifluoroacetate salt hydrate P5749 0 1.0 Z-L-Phe chloromethyl C 9511 0 1.0 (S)-(+)-Camptothecin C 9911 0 1.0 8-Cyclopentyl-1,3-dipropylxanthine C-101 0 1.0 GW9508 G 9797 0 1.0 acetate G-110 0 1.0 (±)-Epinephrine hydrochloride E 4642 0 1.0 Ethosuximide E 7138 0 1.0 E 7881 0 1.0 Disopyramide D 7644 0 1.0 7,7-Dimethyl-(5Z,8Z)-eicosadienoic acid D 8008 0 0.9 CGS-21680 hydrochloride C-141 0 1.0 (+)- C-147 0 1.0 Capsazepine C-191 0 1.0 N-Methyl-beta-carboline-3-carboxamide E-006 0 1.0 C-192 0 1.0 D 8399 0 1.0 4-Imidazolemethanol hydrochloride H 1877 0 1.0 Hexamethonium dichloride H 2138 0 1.0 (R,R)-cis-Diethyl tetrahydro-2,8-chrysenediol D 8690 0 1.0 CGP 20712A methanesulfonate C-231 0 1.0 UCL 2077 U6758 0 1.0 17alpha-hydroxyprogesterone H 5752 0 1.0 Dipropyldopamine hydrobromide D-031 0 1.0 Fenofibrate F 6020 0 1.1 D-ribofuranosylbenzimidazole D 1916 0 1.0 L- hydrochloride H 8125 0 1.1 Forskolin F 6886 0 1.1 Hydroquinone H 9003 0 1.1 R(+)-Butylindazone D-129 0 1.0 F-100 0 1.0 N,N-Dipropyl-5-carboxamidotryptamine maleate D-132 0 1.1 N-(3,3-Diphenylpropyl)glycinamide D 8816 0 1.1 (-)-alpha-Methylnorepinephrine D 5290 0 1.0 Anisotropine methyl bromide A5181 0 1.1 R-(+)-7-Hydroxy-DPAT hydrobromide H-168 0 1.1 Vanillic acid diethylamide E 0137 0 1.0 CPNQ C 3118 0 1.0 Daphnetin D 5564 0 1.1 L-Canavanine C 1625 0 1.1 2',3'-dideoxycytidine D 5782 0 1.0 Cambinol C0494 0 1.0 GBR-12935 dihydrochloride G 9659 0 1.0

3 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( E 1279 0 1.1 ET-18-OCH3 E 1779 0 1.0 5'-N-Ethylcarboxamidoadenosine E 2387 0 1.0 E-64 E 3132 0 1.0 Isoliquiritigenin I 3766 0 0.9 Mecamylamine hydrochloride M 9020 0 0.9 SD-169 S0572 0 1.0 ODQ O 3636 0 1.0 MK-886 M 2692 0 0.9 MDL 105,519 M-216 0 1.0 K 185 K 1888 0 1.0 Meclofenamic acid sodium M 4531 0 1.0 AS605240 A 0233 0 1.0 LY-367,265 L 2411 0 1.0 (-)- tartrate salt N 5260 0 1.0 alpha- hydrochloride L 4376 0 1.0 Valproic acid sodium P 4543 0 1.0 L 9664 0 1.0 T0070907 T 8703 0 1.0 L-655,708 L 9787 0 1.0 NG-Monomethyl-L-arginine acetate M 7033 0 1.0 (±)-3-(3,4-dihydroxyphenyl)-2-methyl-DL- M 7277 0 1.0 SR 59230A oxalate S 8688 0 0.9 SIB 1893 S 9311 0 1.0 hydrochloride R 0758 0 0.9 hydrochloride G 3796 0 1.0 R-103 0 0.9 hydrochloride R-104 0 1.0 3-Tropanyl-indole-3-carboxylate hydrochloride T-104 0 1.0 3-Tropanylindole-3-carboxylate methiodide T-113 0 1.0 5alpha-Pregnan-3alpha-ol-20-one P 8887 0 1.0 Propantheline bromide P 8891 0 1.0 5HPP-33 H 9415 0 0.9 Tyrphostin A9 T-182 0 1.0 hemifumarate R-134 0 1.0 hydrochloride T 1516 0 1.0 Paromomycin sulfate P 9297 0 1.0 Procaine hydrochloride P 9879 0 1.0 Tyrphostin AG 490 T 3434 0 1.0 Org 27569 O8014 0 1.0 TTNPB T 3757 0 1.0 (CDX) B9061 0 1.0 Tyrphostin AG 1478 T 4182 0 1.0 (±)-gamma-Vinyl GABA V 8261 0 1.0 PPADS P-178 0 1.0 Bay 11-7082 B 5556 0 1.0 NSC348884 hydrate N 3414 0 1.0 Albuterol hemisulfate S 5013 0 1.0 (±)-alpha-Lipoic Acid T 5625 0 1.0 DL- T 6031 0 1.0 hydrochloride T 6154 0 1.0 SMER28 S 8197 0 1.0 WIN 62,577 W-104 0 1.0 SR 2640 S 7690 0 1.0 AC-55649 A9480 0 1.0 SKF 96365 S 7809 0 1.0 SCH 58261 S4568 0 1.0 Q 3504 0 1.0 LP 12 hydrochloride hydrate L 3169 0 1.0 Tyrphostin 23 T 7165 0 1.0 Pifithrin-mu P0122 0 1.0 Cortexolone R 0500 0 1.0 1-[2-(Trifluoromethyl)phenyl] T 7313 0 1.0 hydrochloride Y 3125 0 1.1 CV-3988 C 7238 0 1.0 YC-1 Y-102 0 1.0 hydrochloride C 1119 0 1.0 (-)- methyl bromide S 8502 0 1.0

4 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( S(3)3p3Bromotetramisole&oxalate 190047 0 0.9 (±)- maleate B 2390 0 0.9 Azelaic&acid 246379 0 0.9 &hydrochloride 246557 0 1.0 C salt C 3270 0 0.9 &sulfate A&0257 0 0.9 BWB70C B 4558 0 1.0 Bepridil hydrochloride B 5016 0 1.0 (+)-Brompheniramine maleate B 5275 0 1.0 Amiodarone hydrochloride A 8423 0 0.9 3-aminobenzamide A 0788 0 1.0 hydrochloride B 7148 0 1.0 Altretamine A 8723 0 0.9 Cephalothin sodium C 4520 0 1.0 N-(4-Amino-2-chlorophenyl)phthalimide A 9345 0 1.0 N-Acetyl-5-hydroxytryptamine A 1824 0 1.0 BRL 50481 B 0936 0 1.0 Adenosine 3',5'-cyclic monophosphate A 9501 0 0.9 hydrochloride C 5040 0 1.0 (±)-HA-966 A 9699 0 1.0 hydrochloride B-016 0 1.0 Roscovitine R 7772 0 1.0 1,3-Dipropyl-8-p-sulfophenylxanthine A-022 0 1.0 &hydrochloride A&3539 0 1.0 DAPH D 3943 0 1.0 BU224 hydrochloride B-154 0 1.0 N-(2-[4-(4-Chlorophenyl)piperazin-1-yl]ethyl)-3-methoxybenzamide C 7230 0 1.0 Psora-4 P 9872 0 1.0 AS-252424 A8981 0 1.0 PK 11195 C 0424 0 1.0 Acetylthiocholine chloride A 5626 0 1.0 Cantharidin C 7632 0 1.0 Chelidamic acid C 8011 0 1.0 CGP-7930 C 0862 0 1.0 L- C 1159 0 1.0 hydrobromide A 6134 0 1.0 A-315456 A 6351 0 1.0 (+)- Hydrate C 1251 0 1.0 Cephradine C 8395 0 1.0 Actinonin A&6671 0 1.1 hydrochloride C 1671 0 1.0 C 2505 0 1.0 Centrophenoxine hydrochloride C 8773 0 1.0 p-Benzoquinone B 1266 0 1.0 TBBz T6951 0 1.0 CPCCOEt C 9611 0 1.0 Bromoenol lactone B 1552 0 1.0 B 2009 0 1.0 ML-7 I 2764 31 0.9 M 8046 31 0.9 (±)- I 2765 31 0.9 L-alpha-Methyl-p- M 8131 31 0.9 SB-215505 S 1068 31 0.9 Isotharine mesylate I 3639 31 0.9 hydrochloride M 9125 31 1.0 methanesulfonate N-156 31 1.0 3-Isobutyl-1-methylxanthine I 5879 31 0.9 1-(5-Isoquinolinylsulfonyl)-2-methylpiperazine dihydrochloride I 7016 31 0.9 (+)-MK-801 hydrogen maleate M-107 31 1.0 N-Oleoylethanolamine O 0383 31 1.0 O 0877 31 1.0 Stevioside S3572 31 1.0 Ethopropazine hydrochloride E5406 31 1.0 hydrochloride I-120 31 1.0 Mevastatin M 2537 31 0.9 Oxotremorine methiodide O-100 31 1.0 N 0630 31 0.9 3-(1H-Imidazol-4-yl)propyl di(p-fluorophenyl)methyl ether hydrochloride I-160 31 1.0

5 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( (±)-Octoclothepin maleate O-111 31 1.0 isethionate P 0547 31 1.0 O-Phospho-L- P 0878 31 1.0 G8761 31 0.9 L-162,313 L 1415 31 1.0 hydrochloride P 3510 31 1.0 LY-310,762 hydrochloride L 2536 31 1.0 Lomefloxacin hydrochloride L 2906 31 1.0 6-Methyl-2-(phenylethynyl)pyridine hydrochloride M 5435 31 1.0 (±)-Methoxyverapamil hydrochloride M 5644 31 1.0 Nifedipine N 7634 31 1.0 7-Nitroindazole N 7778 31 1.0 CI-976 C3743 31 1.0 GW405833 hydrochloride G1421 31 1.1 2-methoxyestradiol M 6383 31 1.0 alpha,beta-Methylene adenosine 5'-triphosphate dilithium M 6517 31 1.0 hydrochloride P 4670 31 1.0 5alpha-Pregnan-3alpha-ol-11,20-dione P 5052 31 1.0 PNU-282987 P6499 31 1.0 (-)-Tetramisole hydrochloride L 9756 31 1.0 Piroxicam P 5654 31 1.0 3-n-Propylxanthine P 5679 31 1.0 hydrochloride P 6126 31 1.0 P 6402 31 1.0 C 9847 31 1.0 N6-Cyclohexyladenosine C 9901 31 1.0 hydrochloride E 3263 31 1.0 OXA-22 C-011 31 1.0 8-Cyclopentyl-1,3-dimethylxanthine C-102 31 1.0 Disopyramide phosphate D 6035 31 1.0 sodium D 6899 31 1.0 Glipizide G-117 31 1.0 Staurosporine aglycone S 3939 31 1.0 EGTA E 4378 31 1.0 dihydrochloride C3618 31 1.0 Cilnidipine C1493 31 1.0 Y-27632 dihydrochloride Y 0503 31 1.0 Dephostatin D 8065 31 1.0 MHPG H 1377 31 1.0 2-Chloro-2-deoxy-D-glucose C-203 31 1.0 F 0778 31 1.0 CNQX disodium C-239 31 1.0 CX 546 C-271 31 1.0 D 9035 31 1.0 DL-threo-beta-hydroxyaspartic acid H 2775 31 1.0 3,4-Dihydroxyphenylacetic acid D 9128 31 1.0 Chloride L 4408 31 1.0 H 4759 31 1.0 SB 218795 S 8817 31 1.0 Hispidin H 5257 31 1.0 WB-4101 hydrochloride D 0411 31 1.0 Fluvoxamine maleate F 2802 31 1.0 Dihydroouabain D 0670 31 1.0 hydrochloride D 0676 31 1.0 p-Fluoro-L- F 4646 31 1.0 Doxycycline hydrochloride D 9891 31 1.0 6,7-ADTN hydrobromide D-002 31 1.0 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole H 6036 31 1.0 Harmane H 7258 31 1.0 Debrisoquin sulfate D 1306 31 1.1 5-Fluorouracil F 6627 31 1.0 R(+)-SCH-23390 hydrochloride D-054 31 1.0 Diltiazem hydrochloride D 2521 31 1.1 hydrobromide monohydrate D 2531 31 1.1 dihydrochloride F 8257 31 1.0 hydrochloride D 3630 31 1.0 5-fluoro-5'- F 8791 31 1.0 F 9397 31 1.0

6 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( L- H 9002 31 1.1 BU99006 B 8433 31 0.9 D126608 31 1.0 MNS M 7445 31 1.0 hydrochloride H 9523 31 1.0 L-165,041 L 2167 31 1.0 (S)-3,5- D 3689 31 1.1 hydrochloride F 9427 31 1.1 F 9552 31 1.1 Felodipine F 9677 31 1.0 hydrochloride D 3900 31 1.0 cis-(Z)-Flupenthixol dihydrochloride F-114 31 1.0 F-124 31 1.0 FPL 64176 F-131 31 1.0 D(-)-2-Amino-5-phosphonopentanoic acid A 8054 31 1.1 3,7-Dimethyl-1-propargylxanthine D-134 31 1.0 5,7-Dichlorokynurenic acid D-138 31 1.0 , R(-)-alpha-methyl-, dihydrochloride H-128 31 1.0 GW2974 G 0668 31 1.0 L- hydrochloride G 2128 31 1.0 2,3-Dimethoxy-1,4-naphthoquinone D 5439 31 1.0 I 0157 31 1.0 Venlafaxine hydrochloride V7264 31 1.1 Imidazole-4-acetic acid hydrochloride I 0375 31 1.0 Imazodan I 0782 31 1.0 DM 235 D 5689 31 1.1 GW5074 G 6416 31 1.0 G 6649 31 1.0 1,1-Dimethyl-4-phenyl-piperazinium iodide D 5891 31 1.0 Iodoacetamide I 1149 31 1.1 Idarubicin I 1656 31 0.8 GR 55562 dihydrobromide G 0419 31 1.0 sulfate P 6777 31 0.9 maleate P 6902 31 0.9 SKF 89976A hydrochloride S 9066 31 0.9 1-(1-Naphthyl)piperazine hydrochloride S-003 31 1.0 Taxol T 7402 31 0.9 Retinoic acid R 2625 31 0.9 dihydrochloride P 7412 31 1.0 hydrochloride P 7791 31 0.9 L-Beta-threo-benzyl-aspartate B6436 31 1.0 SQ 22536 S-153 31 1.0 R 8875 31 1.0 ARP 101 A8356 31 0.9 hydrochloride P 8511 31 1.0 Ro 8-4304 R 8900 31 0.9 U-75302 U 1508 31 0.9 maleate P 9233 31 1.0 Prilocaine hydrochloride P 9547 31 1.0 Pyridostigmine bromide P 9797 31 1.0 Tolazamide T 2408 31 1.0 5'-diphosphate sodium U 4125 31 1.0 U-69593 U-103 31 1.0 (-)-Scopolamine methyl nitrate S 2250 31 1.0 P-118 31 1.0 IC 261 I 0658 31 1.0 U0126 U-120 31 1.0 tetrahydrochloride S 2876 31 1.0 A-134974 dihydrochloride hydrate A2846 31 0.9 SC 19220 S 3065 31 1.0 PD 98,059 P-215 31 1.0 Tyrphostin AG 835 T 5568 31 1.0 AMG 9810 A 2731 31 1.0 from Penicillium funiculosum W 1628 31 1.0 sulfate Q 0875 31 1.0 BAY 61-3606 hydrochloride hydrate B 9685 31 1.0 N,N,N-trimethyl-1-(4-trans-stilbenoxy)-2-propylammonium iodide T 6692 31 1.0 (-)- S 7771 31 1.0

7 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( (-)-Scopolamine,n-Butyl-, bromide S 7882 31 1.0 , 6-nitro-, maleate Q-109 31 1.0 dihydrochloride Q-110 31 1.0 (±)- dihydrochloride Q-111 31 1.0 Aprindine hydrochloride A 7606 31 1.0 (±)-Sulpiride S 8010 31 1.1 YS-035 hydrochloride Y-101 31 1.0 Z 0878 31 1.0 hydrochloride O 7389 31 1.0 DL3alpha3Methyl3p3tyrosine 120693 31 0.9 3-Bromo-7-nitroindazole B 2050 31 0.9 53Aminovaleric&acid&hydrochloride 194336 31 0.9 N-Bromoacetamide B 2377 31 0.9 Benzamil hydrochloride B 2417 31 1.0 L-2-aminoadipic acid A 7275 31 0.9 (±)- A 7655 31 0.9 acetate C 3412 31 0.9 5-Bromo-2'-deoxyuridine B 5002 31 1.0 GR 113808 G 5918 31 1.0 C 4024 31 1.0 (±)- B 5399 31 0.9 B 7005 31 1.0 4-Amino-1,8-naphthalimide A 0966 31 1.0 Carbachol C 4382 31 0.9 Adenosine A 9251 31 0.9 Astaxanthin A 9335 31 1.0 &hydrochloride A&1260 31 0.9 N-arachidonylglycine A 1977 31 1.0 BP 897 B 9308 31 1.0 3'3Azido33'3deoxythymidine A&2169 31 0.9 (E)-5-(2-Bromovinyl)-2'-deoxyuridine B 9647 31 1.0 5-azacytidine A 2385 31 0.9 Bromoacetyl menthane B-015 31 1.0 A 3145 31 1.0 hydrochloride B-019 31 1.0 Paroxetine hydrochloride hemihydrate (MW = 374.83) P 9623 31 0.9 SB 204741 S 0693 31 0.9 hydrochloride B-102 31 1.0 1-Aminobenzotriazole A 3940 31 1.0 3-Amino-1-propanesulfonic acid sodium A 4147 31 1.0 hydrochloride C 6022 31 1.0 propargyl ester hydrobromide A-140 31 1.0 O3(Carboxymethyl)hydroxylamine&hemiHCl A&4508 31 1.0 Sandoz 58-035 S 9318 31 1.0 3-Morpholinosydnonimine hydrochloride M5793 31 1.0 Diacylglycerol kinase inhibitor I D 5919 31 1.0 (±)323Amino373phosphonoheptanoic&acid A&5157 31 1.0 Ciprofibrate C 0330 31 1.0 6-Chloromelatonin C 0331 31 1.0 C 0737 31 1.0 N6-2-(4-Aminophenyl)ethyladenosine A-202 31 1.0 C 7971 31 1.0 gamma-Acetylinic GABA A-230 31 1.0 Alloxazine A-242 31 1.0 Acetazolamide A 6011 31 1.0 GR 46611 G 8543 31 1.0 Chlorpropamide C 1290 31 1.0 cis-Azetidine-2,4- A-243 31 1.1 trans-Azetidine-2,4-dicarboxylic acid A-244 31 1.0 AIDA A-254 31 1.0 monohydrate I 0160 31 1.0 ATPA A-263 31 1.0 ARL 67156 trisodium salt A-265 31 1.0 C 8759 31 1.0 &hydrate A&6770 31 1.0 hydrochloride A 6976 31 1.0 Aminoguanidine hemisulfate A 7009 31 1.0 3-Aminopropylphosphonic acid A 7162 31 1.0

8 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( 2,3-Butanedione monoxime B 0753 31 1.1 Pyrocatechol C 9510 31 1.0 Colchicine C 9754 31 1.0 1-Methylimidazole M 8878 32 0.9 (±)- I 4883 32 1.0 hydrochloride M 9292 32 1.0 Nitrendipine N-144 32 0.9 BW 723C86 B175 32 0.9 SB 242084 dihydrochloride hydrate S 8061 32 0.9 dihydrochloride N-158 32 1.0 F 0430 32 1.0 (±)- chloride M-104 32 1.0 DFB D 0943 32 1.0 I 8898 32 1.0 maleate M-137 32 1.0 Olomoucine O 0886 32 0.9 O 3011 32 1.0 Indatraline hydrochloride I-119 32 0.9 S15535 S 5321 32 1.0 hydrochloride O 3752 32 0.9 Mexiletene hydrochloride M 2727 32 1.0 IB-MECA I-146 32 0.9 I18008 32 1.0 Nomifensine maleate N 1530 32 1.0 N-Methyl-D-aspartic acid M 3262 32 1.0 JL-18 J-102 32 1.0 nor- dihydrochloride N 1771 32 1.0 Niclosamide N 3510 32 1.0 M 3668 32 1.0 L-701,324 L 0258 32 1.0 Nordihydroguaiaretic acid from Larrea divaricata (creosote bush) N 5023 32 1.0 3-alpha,21-Dihydroxy-5-alpha-pregnan-20-one P 2016 32 1.0 (±)-alpha-Methyl-4-carboxyphenylglycine M 4796 32 1.0 PRE-084 P 2607 32 1.0 NG-Nitro-L-arginine N 5501 32 1.0 NG-Nitro-L-arginine methyl ester hydrochloride N 5751 32 1.0 Mibefradil dihydrochloride M 5441 32 1.0 N6-Methyladenosine M 5501 32 1.0 NU2058 N 4286 32 1.0 Leflunomide L 5025 32 1.0 Lidocaine N-ethyl bromide quaternary salt L 5783 32 1.0 4-Amidinophenylmethanesulfonyl fluoride hydrochloride A 6664 32 1.0 NS 521 oxalate N 7904 32 1.0 LE 300 L 8401 32 1.0 L 8533 32 1.0 6-Nitroso-1,2-benzopyrone N 8403 32 1.0 N 8534 32 1.0 hydrochloride P 4651 32 1.0 Cysteamine hydrochloride M 6500 32 1.0 hydrochloride M 6524 32 1.0 O-Methylserotonin hydrochloride M 6628 32 1.0 Phosphomycin disodium P 5396 32 1.0 Norcantharidin N 8784 32 1.0 hydrchloride N 9007 32 1.0 Pentylenetetrazole P 6500 32 1.0 (+)-Pilocarpine hydrochloride P 6503 32 1.0 2,6-Difluoro-4-[2-(phenylsulfonylamino)ethylthio]phenoxyacetamide D 8941 32 1.0 SB 415286 S 3567 32 1.0 S-Ethylisothiourea hydrobromide E 3149 32 1.0 hydrochloride E 5156 32 1.0 Ebselen E 3520 32 1.0 Diethylenetriaminepentaacetic acid D 6518 32 1.0 R-(-)-Desmethyldeprenyl hydrochloride D 6940 32 1.0 (±)- H 0131 32 1.0 6-Hydroxymelatonin H 0627 32 1.0 Hexamethonium bromide H 0879 32 1.0 (-)-Physostigmine E 8375 32 1.0 NBI 27914 N 3911 32 1.0

9 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( 8-(3-Chlorostyryl) C-197 32 1.0 Imperatorin I6659 32 1.0 CGS-15943 C-199 32 1.0 Fusidic acid sodium F 0881 32 1.0 S-(+)- hydrochloride F 1553 32 1.0 DCEBIO D 9190 32 1.0 1-Deoxynojirimycin hydrochloride D 9305 32 1.0 D 9766 32 1.0 Furegrelate sodium F 3764 32 1.0 F 4381 32 1.0 dihydrochloride F 4765 32 0.9 R(-)-2,10,11-Trihydroxyaporphine hybrobromide D-029 32 1.0 2',3'-didehydro-3'-deoxythymidine D 1413 32 1.0 D 1414 32 1.0 L-3,4-Dihydroxyphenylalanine methyl ester hydrochloride D 1507 32 1.0 GBR-12909 dihydrochloride D-052 32 1.0 MHPG sulfate potassium H 8759 32 1.0 (±)-2,3-Dichloro-alpha-methylbenzylamine hydrochloride D-103 32 1.0 5-Hydroxyindolacetic acid H 8876 32 1.0 C3993 32 1.0 4-DAMP methiodide D-104 32 1.1 succinate D 3775 32 1.1 N^G,N^G-Dimethylarginine hydrochloride D 4268 32 1.0 Clodronic acid D 4434 32 1.0 N- D5676 32 1.0 Hexahydro-sila-difenidol hydrochloride, p-fluoro analog H-127 32 1.0 AL-8810 A 3846 32 1.0 sodium D 4505 32 1.1 S(-)- P-152 32 1.0 Ganaxolone G 7795 32 1.0 G 2536 32 1.0 Sematilide monohydrochloride monohydrate S0323 32 1.1 Epibestatin hydrochloride E 0381 32 1.0 Bezafibrate B 7273 32 1.1 5,5-Dimethyl-1-pyrroline-N-oxide D 5766 32 1.0 N-Methyldopamine hydrochloride D 5886 32 1.0 Gallamine triethiodide G 8134 32 1.0 Ipratropium bromide I 1637 32 1.0 hydrochloride E 1896 32 1.0 SU 4312 S 8567 32 0.9 BRL 52537 hydrochloride B 5559 32 0.9 Tetracaine hydrochloride T 7508 32 0.9 Phorbol 12-myristate 13-acetate P 8139 32 1.0 NU6027 N 4411 32 0.9 D-609 potassium T 8543 32 1.0 disodium R 7385 32 1.0 Picotamide P 8477 32 1.0 pyrrolidinedithiocarbamate P 8765 32 1.0 Tamoxifen citrate T 9262 32 0.9 Ibandronate sodium I5784 32 1.0 (±)-cis-Piperidine-2,3-dicarboxylic acid P 8782 32 1.0 Tetramisole hydrochloride T 1512 32 1.0 S(+)- L-tartrate R-121 32 1.0 Procainamide hydrochloride P 9391 32 1.0 Tetradecylthioacetic acid T 1698 32 1.0 (±)- hydrochloride S 0278 32 1.0 SB-366791 S 0441 32 1.0 (±)- S 0752 32 1.0 SKF 95282 dimaleate S 5317 32 1.0 4-Imidazoleacrylic acid U 7500 32 1.0 N2-Ethyl-2'- N 3289 32 1.0 hemisulfate T 2528 32 1.0 Triamterene T 4143 32 1.0 Auranofin A 6733 32 1.0 DL-Stearoylcarnitine chloride S 2381 32 1.0 BF-170 hydrochloride B 4311 32 1.0 Tyrphostin AG 494 T 4318 32 0.9 A3 hydrochloride A 1980 32 1.0

10 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( trihydrochloride S 2501 32 1.0 U-99194A maleate U-116 32 1.0 sulfate salt V 1377 32 1.0 V 6383 32 1.0 Phenamil methanesulfonate P-203 32 1.0 Tyrphostin AG 555 T 4818 32 1.0 (±)-PD 128,907 hydrochloride P-216 32 1.0 Stattic S7947 32 1.0 ICI 63,137 I 8283 32 1.0 Sanguinarine chloride S 5890 32 1.0 LY2183240 L 9044 32 1.0 N-Succinyl-L- S 6633 32 1.0 SU 6656 S 9692 32 1.0 P63204 32 1.0 T 6376 32 1.0 Quinine sulfate Q 1250 32 1.0 S(-)- maleate T 6394 32 1.0 (+)- Q 2128 32 1.0 IRAK-1/4 Inhibitor I I 5409 32 1.1 Ara-G hydrate A 4233 32 1.0 hydrochloride X 1251 32 1.0 amine congener X-103 32 1.0 Tyrphostin AG 112 T 6943 32 1.0 (-)-Quinpirole hydrochloride Q-102 32 1.0 Tyrphostin 1 T 7040 32 1.0 O6-benzylguanine B 2292 32 0.9 L-Buthionine-sulfoximine B 2515 32 1.0 53Fluoroindole323carboxylic&acid 265128 32 1.0 Chlormethiazole hydrochloride C1240 32 1.0 CGP-74514A hydrochloride C 3353 32 0.9 S3(43Nitrobenzyl)363thioguanosine 861669 32 0.9 1-Aminocyclopropanecarboxylic acid hydrochloride A 0430 32 1.0 D-Cycloserine C 3909 32 0.9 4-(2-Aminoethyl)benzenesulfonyl fluoride hydrochloride A 8456 32 0.9 Org&24598&lithium&salt O7639 32 0.9 Benzamidine hydrochloride B 6506 32 1.0 hydrochloride C 4542 32 1.0 ðylenediamine A&1755 32 0.9 Ro 20-1724 B 8279 32 1.0 Chlorothiazide C 4911 32 0.9 Bethanechol chloride C 5259 32 1.0 Acetyl-beta-methylcholine chloride A 2251 32 0.9 5-(N-Ethyl-N-isopropyl)amiloride A 3085 32 1.0 (±)-AMT hydrochloride A 9834 32 1.0 Ceftriaxone sodium C 5793 32 1.0 HEMADO H3288 32 1.0 (-)-Cotinine C 5923 32 1.0 Clotrimazole C 6019 32 1.0 YM&976 Y&4877 32 1.0 AA-861 A 3711 32 1.0 9-Amino-1,2,3,4-tetrahydroacridine hydrochloride A 3773 32 1.0 Bromoacetylcholine bromide B-121 32 1.0 R(+)-6-Bromo-APB hydrobromide B-135 32 1.0 S(-)-Atenolol A-143 32 1.0 53(N,N3Dimethyl)amiloride&hydrochloride A&4562 32 1.0 A 4638 32 1.0 Acyclovir A 4669 32 1.0 BRL 54443 maleate B-173 32 1.0 (±)-2-Amino-3-phosphonopropionic acid A 4910 32 1.0 Supercinnamaldehyde S3322 32 0.9 DL-Cycloserine C 7005 32 1.0 1-Amino-1-cyclohexanecarboxylic acid hydrochloride A-162 32 1.0 McN-A-343 C 7041 32 1.0 Cystamine dihydrochloride C 7255 32 1.0 SB 200646 hydrochloride S 0568 32 1.0 Caffeine C 0750 32 1.0 Acetohexamide A-178 32 1.0 AB-MECA A-236 32 1.0

11 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( ML-9 C 1172 32 1.0 2-Hydroxysaclofen A 6566 32 1.0 hydrochloride C 8138 32 1.0 CBIQ C 6617 32 1.0 Caffeic acid phenethyl ester C 8221 32 1.0 DSP-4 hydrochloride C 8417 32 1.0 CB 1954 C 2235 32 1.0 4-Aminobenzamidine dihydrochloride A 7148 32 1.0 1-benzoyl-5-methoxy-2-methylindole-3-acetic acid B 1183 32 1.0 8-Bromo-cGMP sodium B 1381 32 1.0 (±)-Isoproterenol hydrochloride I 5627 33 1.0 R(-)-Me5 M 0814 33 1.0 phosphate I 7627 33 0.9 (-)-MK-801 hydrogen maleate M-108 33 0.9 Cibenzoline succinate C1618 33 0.9 L-alpha-Methyl DOPA M-129 33 1.0 4-Methylpyrazole hydrochloride M 1387 33 0.9 Chloride O 2881 33 0.9 MRS 1523 M 1809 33 1.0 ZM 39923 hydrochloride Z 4626 33 0.9 SB-525334 S 8822 33 1.0 hydrochloride M 2525 33 0.9 SB 216763 S 3442 33 1.0 dihydrobromide I-135 33 1.0 1,5-Isoquinolinediol I-138 33 0.9 Nimesulide N 1016 33 1.0 Molsidomine M 2901 33 1.0 BBMP B7936 33 0.9 Mizoribine M 3047 33 1.0 MG 624 M 3184 33 1.0 alpha-Methyl-DL-tyrosine methyl ester hydrochloride M 3281 33 1.0 K 0250 33 1.0 tris salt K 1136 33 1.0 fumarate K 2628 33 1.0 NAN-190 hydrobromide N 3529 33 1.0 P 1793 33 1.0 U-73343 U 6881 33 1.0 S-Nitrosoglutathione N 4148 33 1.0 M 4659 33 1.0 1-Methylhistamine dihydrochloride M 4910 33 1.0 1,3-Dimethyl-8-phenylxanthine P 2278 33 1.0 S-Methyl-L-thiocitrulline acetate M 5171 33 1.0 M 5250 33 1.0 L- sulfoximine M 5379 33 1.0 hydrochloride N 5504 33 1.0 3-Nitropropionic acid N 5636 33 1.0 (±)- hydrochloride N 7127 33 1.0 1-Phenyl-3-(2-thiazolyl)-2-thiourea P 4015 33 1.0 Podophyllotoxin P 4405 33 1.0 VER-3323 hemifumarate salt V 1889 33 1.0 Lidocaine hydrochloride L 5647 33 1.0 L-687,384 hydrochloride L 8539 33 1.0 Praziquantel P 4668 33 1.0 LY-294,002 hydrochloride L 9908 33 1.0 succinate L-106 33 1.0 GR 127935 hydrochloride hydrate G 5793 33 0.9 Edrophonium chloride E 3256 33 1.0 Ellipticine E 3380 33 1.0 hydrochloride G-002 33 1.0 hydrochloride G-007 33 1.0 hydrochloride D 6140 33 1.0 hydrobromide G-019 33 1.0 2,2'-Bipyridyl D 7505 33 1.0 CGS-12066A maleate C-106 33 1.0 N-Ethylmaleimide E 3876 33 1.0 2-Cyclooctyl-2-hydroxyethylamine hydrochloride C-108 33 1.0 5-Carboxamidotryptamine maleate C-117 33 1.0 (±)-CGP-12177A hydrochloride C-125 33 1.0

12 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( DL-Homatropine hydrobromide H 0126 33 1.0 3,4-Dichloroisocoumarin D 7910 33 1.0 beta- E 8875 33 1.0 Phenserine P0111 33 1.0 Hypotaurine H 1384 33 1.0 H 1512 33 1.0 5'-diphosphocholine sodium salt hydrate C0256 33 1.0 hydrochloride H 1753 33 1.0 S-(-)- hydrochloride E-101 33 1.0 hydrobromide F 1016 33 1.0 hydrochloride H 3132 33 1.0 Dantrolene sodium D 9175 33 1.0 H 4001 33 1.0 R(-)-Propylnorapomorphine hydrochloride D-027 33 1.0 Histamine dihydrochloride H 7250 33 1.0 JS-K J 4137 33 1.1 Retinoic acid p-hydroxyanilide H 7779 33 1.0 hydrochloride F 6145 33 1.0 Genipin G4796 33 1.0 1,4-Dideoxy-1,4-imino-D-arabinitol D 1542 33 1.0 F 6513 33 1.1 dihydrate E 7906 33 1.0 bromide F 6800 33 0.9 R(-)-N-Allylnorapomorphine hydrobromide D-042 33 1.1 (±)-SKF-38393 hydrochloride D-047 33 1.0 Hydroxyurea H 8627 33 1.0 (+)- H 8645 33 1.0 (±)-DOI hydrochloride D-101 33 1.0 F 6889 33 1.0 FSCPX F 7927 33 1.0 methanesulfonate D 2763 33 1.0 NS8593 hydrochloride N 2538 33 1.0 2,3-Butanedione D 3634 33 1.0 maleate F 8927 33 1.0 1,3-Dipropyl-7-methylxanthine D-108 33 1.1 D-122 33 1.0 E 2031 33 1.0 5-Hydroxy-L-tryptophan H 9772 33 1.0 N-Methyl-1-deoxynojirimycin M 1777 33 1.0 Fluoxetine hydrochloride F-132 33 1.0 Dihydro-beta-erythroidine hydrobromide D-149 33 1.0 hydrochloride D 5294 33 1.0 1,7-Dimethylxanthine D 5385 33 1.0 L- G 3126 33 1.0 DL-alpha-Difluoromethylornithine hydrochloride D-193 33 1.0 ATPO A 7845 33 1.0 NSC 95397 N 1786 33 1.0 E 0516 33 1.0 alpha-Guanidinoglutaric acid G 7788 33 1.0 HA-100 I 1392 33 1.0 Phosphonoacetic acid P 6909 33 0.9 hydrochloride R 2530 33 1.0 R 5010 33 1.0 dihydrochloride P 7505 33 0.9 PAPP S-009 33 0.9 SR-95531 S-106 33 0.9 hydrochloride P 8013 33 1.0 Rottlerin R 5648 33 0.9 Ro 25-6981 hydrochloride R 7150 33 1.0 hydrochloride T 9025 33 0.9 Thapsigargin T 9033 33 1.0 Sepiapterin S-154 33 0.9 A 2729 33 0.9 SB 206553 hydrochloride S-180 33 0.9 (S)- hydrochloride P 8688 33 1.0 RX 821002 hydrochloride R 9525 33 0.9 T 9652 33 0.9 hydrochloride R-101 33 0.9

13 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( hydrochloride T-103 33 1.0 Ro 41-1049 hydrochloride R-107 33 1.0 hydrochloride T 0410 33 0.9 hydrochloride P 8813 33 0.9 6(5H)-Phenanthridinone P 8852 33 1.0 Tetraethylthiuram disulfide T 1132 33 1.0 TCPOBOP T 1443 33 1.0 K114 K 1015 33 1.0 dimaleate P 9178 33 1.0 dihydrochloride T-122 33 1.0 maleate T-123 33 0.9 R-116 33 0.9 Steviol H8664 33 1.0 A-68930 hydrochloride A 8852 33 0.9 hydrochloride T 6950 33 1.0 Theophylline T 1633 33 0.9 1,10- monohydrate P 9375 33 1.0 Ro 04-6790 dihydrochloride R-140 33 1.0 4-Hydroxyphenethylamine hydrochloride T 2879 33 1.0 R(+)-3PPP hydrochloride P-102 33 1.0 S(-)-3PPP hydrochloride P-103 33 1.0 L-765,314 L 3040 33 1.0 UK 14,304 U-104 33 1.0 Tetrabenazine T2952 33 1.0 (+)- hydrochloride N3288 33 1.0 hydrochloride S 2201 33 1.0 U-101958 maleate U-115 33 1.0 Tetrahydrozoline hydrochloride T 4264 33 1.0 1- P-120 33 1.0 SID7969543 S1323 33 1.0 P-126 33 1.0 cDPCP C 0996 33 0.9 (±)- T 4512 33 1.0 SNC80 S 2812 33 1.0 (±)-Verapamil hydrochloride V 4629 33 1.0 hydrochloride T 4680 33 1.0 N-Oleoyldopamine O 2139 33 1.0 L- from Koa hoale seeds M0253 33 1.0 D-Serine S 4250 33 1.0 PD 168,077 maleate P-233 33 1.0 SB 269970 hydrochloride S 7389 33 1.0 hydrochloride S 7395 33 1.0 WAY-100635 maleate W-108 33 1.0 SB 205384 S 7936 33 1.0 Na-p-Tosyl-L- chloromethyl ketone hydrochloride T 7254 33 1.0 chloride C 2932 33 0.9 N-Acetyltryptamine A 7342 33 0.9 Cortisone 21-acetate C 3130 33 0.9 L- A 7762 33 1.0 B 3023 33 0.9 Benazoline oxalate B 4555 33 0.9 Atropine&methyl&nitrate A&0382 33 1.0 Arcaine&sulfate A&0384 33 1.0 ATPO A 7845 33 0.9 Calmidazolium chloride C 3930 33 0.9 SB 202190 S 7067 33 0.9 (±)- (+)bitartrate A 0937 33 0.9 Brefeldin A from Penicillium brefeldianum B 7651 33 0.9 (±)-2-Amino-4-phosphonobutyric acid A 1910 33 1.0 hydrochloride A 0232 33 1.0 Cefazolin sodium C 5020 33 0.9 (±)-p- A 9657 33 1.0 A 9950 33 0.9 CL 316,243 C 5976 33 1.0 7-Chloro-4-hydroxy-2-phenyl-1,8-naphthyridine C 5982 33 1.0 (-)- methbromide, 1(S), 9(R) B-103 33 1.0 10058-F4 F3680 33 1.0 BTCP hydrochloride B-138 33 1.0

14 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( 5'-Amino-5'- p-toluenesulfonate salt A283 33 1.0 GR 79236X G5794 33 1.0 C 6305 33 1.0 Aminobenztropine A-138 33 1.0 (±)-p-Chlorophenylalanine C 6506 33 1.0 diphosphate C 6628 33 1.0 B-HT 933 dihydrochloride B-161 33 1.0 hydrochloride B 5311 33 1.0 Chlorambucil C 0253 33 1.0 L-Arginine A 5006 33 1.0 1-Allyl-3,7-dimethyl-8-p-sulfophenylxanthine A-145 33 1.0 (±)323Amino353phosphonopentanoic&acid A&5282 33 1.0 L-732,138 A 5330 33 0.9 Acetylsalicylic acid A 5376 33 1.0 4-Androsten-4-ol-3,17-dione A 5791 33 1.0 2-(2-Aminoethyl)isothiourea dihydrobromide A 5879 33 1.0 Cyclophosphamide monohydrate C 0768 33 1.0 SKF-89145 hydrobromide S 3316 33 1.0 cis-4-Aminocrotonic acid A-201 33 1.0 N3Acetylprocainamide&HCl A&5909 33 1.0 Amifostine A 5922 33 1.0 A-77636 hydrochloride A-255 33 0.9 Beclomethasone B 0385 33 1.0 Choline bromide C 1754 33 1.0 Carboplatin C 2538 33 1.0 fumarate C 8903 33 1.0 beta-Chloro-L-alanine hydrochloride C 9033 33 1.0 tartrate I 2892 34 0.9 TMPH hydrochloride T5576 34 0.9 Nimodipine N-149 34 0.9 cis(+/-)-8-OH-PBZI hydrobromide P 0618 34 0.9 tartrate L-121 34 1.0 AFMK A 2355 34 1.0 hydrochloride M 9511 34 0.9 M-001 34 0.9 hydrochloride M 0763 34 1.0 NS 2028 N-211 34 1.0 (±)- hydrochloride O 0250 34 1.0 L-N6-(1-Iminoethyl)lysine hydrochloride I 8021 34 0.9 3-Iodo-L-tyrosine I 8250 34 0.9 hydrochloride I 9531 34 1.0 Oleic Acid O 1008 34 0.9 Moxonidine hydrochloride M 1559 34 0.9 Ouabain O 3125 34 1.0 ICI 204,448 hydrochloride I-122 34 1.0 p-MPPI hydrochloride M-204 34 0.9 Metaproterenol hemisulfate M 2398 34 0.9 TG003 T 5575 34 0.9 Oxotremorine sesquifumarate salt O 9126 34 0.9 8-Methoxymethyl-3-isobutyl-1-methylxanthine M 2547 34 0.9 Trandolapril T4827 34 1.0 Oxaprozin O 9637 34 1.0 hydrochloride M 1818 34 0.9 p-MPPF dihydrochloride M-226 34 0.9 L8668 34 1.0 N 1392 34 1.0 JWH-015 J 4252 34 1.0 P 0130 34 0.9 S-Methylisothiourea hemisulfate M 3127 34 1.0 S-(4-Nitrobenzyl)-6-thioinosine N 2255 34 0.9 S-Nitroso-N-acetylpenicillamine N 3398 34 1.0 (±)-Propranolol hydrochloride P 0884 34 1.0 MRS 2179 M 3808 34 1.0 Meloxicam sodium M 3935 34 1.0 Morin M 4008 34 1.0 P 1784 34 1.0 Minoxidil M 4145 34 1.0 L-Glutamic acid, N-phthaloyl- P 1801 34 1.0

15 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( NCS-382 N 4159 34 1.0 loxoprofen L 0664 34 1.0 hydrochloride L 1011 34 1.0 5-Nitro-2-(3-phenylpropylamino)benzoic acid N 4779 34 1.0 AMN082 A6605 34 1.0 Pirfenidone P 2116 34 1.0 Olvanil O 0257 34 1.0 hydrochloride L 4762 34 1.0 Lonidamine L 4900 34 1.0 NADPH tetrasodium N 7505 34 1.0 Thiolactomycin T 9567 34 1.0 (S)-MAP4 hydrochloride M 5560 34 1.0 Palmitoyl-DL-Carnitine chloride P 4509 34 1.0 hydrochloride N 7758 34 1.0 P 5295 34 1.0 hydrochloride N-115 34 1.0 Sertraline hydrochloride S 6319 34 1.0 (±)-AMPA hydrobromide G-017 34 0.9 S-(-)- C1335 34 1.0 (±)-Chloro-APB hydrobromide C-130 34 1.0 P0099 34 1.0 Daidzein D 7802 34 1.0 GR-89696 fumarate G-133 34 1.0 Dicyclomine hydrochloride D 7909 34 1.0 (±) trans-U-50488 methanesulfonate D 8040 34 1.0 1-(m-Chlorophenyl)-biguanide hydrochloride C-144 34 1.0 Estrone E 9750 34 1.0 3-deazaadenosine D 8296 34 1.0 Bendamustine hydrochloride B 5437 34 1.0 erythro-9-(2-Hydroxy-3-nonyl) hydrochloride E-114 34 1.0 hydrochloride C-223 34 1.0 Alinidine A7230 34 1.0 Hydroxytacrine maleate H 3146 34 1.0 mesylate D 9815 34 1.0 DNQX D 0540 34 1.0 R(-)-Apocodeine hydrochloride D-003 34 1.0 1-(4-Hydroxybenzyl)imidazole-2-thiol H 6892 34 1.0 F 6300 34 1.0 SANT-1 S4572 34 1.0 Mephetyl tetrazole M 7945 34 1.1 Icilin I 9532 34 1.0 hydrochloride H 8502 34 1.0 (±)-7-Hydroxy-DPAT hydrobromide H 8653 34 1.0 N,N,N',N'-Tetramethylazodicarboxamide D 3648 34 1.0 BNTX maleate salt hydrate B 8312 34 1.0 N-Methylhistaprodifen dioxalate salt M 7320 34 1.0 5-hydroxydecanoic acid sodium H-135 34 1.0 1-(2-Methoxyphenyl)piperazine hydrochloride S-008 34 0.9 Propionylpromazine hydrochloride P 7780 34 0.9 (±)-6-Chloro-PB hydrobromide S-143 34 0.6 Phloretin P 7912 34 1.0 BIA 2-093 B 5435 34 1.0 (±)-SKF 38393, N-allyl-, hydrobromide S-168 34 0.9 Phenylbutazone P 8386 34 1.0 T 0625 34 0.9 CCT007093 C 9369 34 1.0 Sodium nitroprusside dihydrate S 0501 34 1.0 hydrochloride U-100 34 1.0 Sulindac sulfone S 1438 34 1.0 2-Phenylaminoadenosine P-101 34 1.0 maleate T 3146 34 1.0 (±)-PPHT hydrochloride P-105 34 1.0 SKF 86466 S 1563 34 1.0 (-)-Scopolamine hydrobromide S 1875 34 1.0 S(-)-UH-301 hydrochloride U-108 34 1.0 (-)-trans-(1S,2S)-U-50488 hydrochloride U-111 34 1.0 N-p-Tosyl-L-phenylalanine chloromethyl ketone T 4376 34 1.0 (6R)-5,6,7,8-Tetrahydro-L-biopterin hydrochloride T 4425 34 1.0

16 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( SKF 83959 hydrobromide S 2816 34 1.0 S(+)-PD 128,907 hydrochloride P-183 34 1.0 Spironolactone S 3378 34 1.0 (±)-Vesamicol hydrochloride V-100 34 1.0 hydrochloride T 6050 34 1.0 1-(2-Chlorophenyl)-1-(4-chlorophenyl)-2,2-dichloroethane C 3010 34 0.9 TMB38&hydrochloride 861804 34 0.9 43aminopyridine A&0152 34 0.9 A 8003 34 0.9 Alprenolol hydrochloride A 8676 34 1.0 Bay 11-7085 B 5681 34 0.9 p-Aminoclonidine hydrochloride A 0779 34 0.9 Reserpine R 0875 34 1.0 BW 284c51 A 9013 34 0.9 A 1784 34 0.9 Bestatin hydrochloride B 8385 34 1.0 Cephalexin hydrate C 4895 34 0.9 L(-)-Norepinephrine bitartrate A 9512 34 0.9 BRL 15572 B 9929 34 0.9 Chloroethylclonidine dihydrochloride B-003 34 1.0 Antozoline hydrochloride A 9899 34 0.9 PNU-37883A P 0248 34 1.0 A-129 34 1.0 -1-beta-D-arabinofuranoside hydrochloride C 6645 34 0.9 BRL 37344 sodium B-169 34 1.0 Amiprilose hydrochloride A 4687 34 1.0 hydrochloride A-164 34 1.0 hydrochloride C 7291 34 1.0 Carmustine C 0400 34 1.0 Aconitine A 8001 34 1.0 Citalopram hydrobromide C 7861 34 1.0 hydrochloride C 7897 34 1.0 CP55940 C 1112 34 1.0 Cinoxacin C 8645 34 1.0 Atropine methyl bromide A 6883 34 1.0 BTO-1 B 6311 34 1.0 sodium L-109 35 0.9 Nylidrin hydrochloride N-153 35 0.9 (-)-Isoproterenol hydrochloride I 6504 35 0.9 Indomethacin I 7378 35 1.0 Isoxanthopterin I 7388 35 1.0 Dihydrocapsaicin M 1022 35 1.0 S(+)-Isoproterenol (+)-bitartrate I 8005 35 0.9 Nocodazole M 1404 35 0.9 Melphalan M 2011 35 1.0 m-Iodobenzylguanidine hemisulfate I 9890 35 0.9 Methiothepin mesylate M-149 35 0.9 Nemadipine-A N4163 35 0.9 3-Morpholinosydnonimine hydrochloride M-184 35 1.0 3-Methoxy-morphanin hydrochloride M-187 35 1.0 Methylergonovine maleate M 2776 35 1.0 Sivelestat sodium salt hydrate S 7198 35 0.9 ML 10302 M 7319 35 1.0 Pindolol P 0778 35 1.0 Neostigmine bromide N2001 35 1.0 K 1003 35 1.0 K 1751 35 1.0 (-)-cis-(1S,2R)-U-50488 tartrate U-106 35 1.0 P 1675 35 1.0 LP44 L9793 35 1.0 Rufinamide R 8404 35 1.0 Nalidixic acid sodium N 4382 35 1.0 CyPPA C5493 35 1.0 beta-Lapachone L 2037 35 1.0 Lamotrigine L 3791 35 1.0 hydrochloride N 7261 35 1.0 Metrazoline oxalate M 5685 35 1.0 GW9662 M 6191 35 1.0

17 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( Se-(methyl)selenocysteine hydrochloride M 6680 35 1.0 Ro 90-7501 R 0529 35 0.6 (-)-Epinephrine bitartrate E 4375 35 1.0 DBO-83 D 7938 35 1.0 Tamoxifen T 5648 35 1.0 Tocainide hydrochloride T0202 35 1.0 AS 604850 A 0231 35 1.0 Fiduxosin hydrochloride F 4303 35 0.9 Diphenyleneiodonium chloride D 2926 35 1.0 N 2288 35 1.0 R-(+)-8-Hydroxy-DPAT hydrobromide H-140 35 1.0 hydrochloride D 4526 35 1.0 Tomoxetine T 7947 35 0.9 1,4-PBIT dihydrobromide P 8352 35 0.9 THIP hydrochloride T-101 35 1.0 XCT790 X4753 35 1.0 Tolbutamide T 0891 35 1.0 Tetraisopropyl pyrophosphoramide T 1505 35 1.0 TPMPA T-200 35 1.0 (E)-4-amino-2-butenoic acid T 1694 35 1.0 U-74389G maleate U 5882 35 0.9 Urapidil, 5-Methyl- U-101 35 1.0 hydrochloride T 2896 35 1.0 Enalaprilat dihydrate E9658 35 1.0 U-62066 U-105 35 1.0 CGP 57380 C0993 35 1.0 T 4500 35 1.0 Pinacidil P-154 35 1.0 SKF 75670 hydrobromide S 2941 35 1.0 hydrochloride from Streptomyces orientalis V 8138 35 1.0 Phenylbenzene-omega-phosphono-alpha- P-204 35 1.0 Thio-NADP sodium T 5515 35 1.0 Betaine hydrochloride B 3501 35 0.9 Betaine chloride B 3650 35 0.9 hydrochloride A 8404 35 0.9 Ancitabine hydrochloride A 8598 35 1.0 L-azetidine-2-carboxylic acid A 0760 35 0.9 Chlorzoxazone C 4397 35 0.9 Aminoguanidine hydrochloride A 8835 35 0.9 L-Cysteinesulfinic Acid C 4418 35 0.9 B 7777 35 0.9 S-(p-Azidophenacyl) A 1782 35 0.9 8-Bromo-cAMP sodium B 7880 35 0.9 Benztropine mesylate B 8262 35 1.0 5-(N,N-hexamethylene)amiloride A 9561 35 0.9 4-Chloromercuribenzoic acid C 5913 35 1.0 (±)-Bay K 8644 B-112 35 0.9 BMY 7378 dihydrochloride B-134 35 1.0 T-112 35 1.0 Cefmetazole sodium C 6048 35 1.0 Clofibrate C 6643 35 1.0 (±)-Butaclamol hydrochloride B-168 35 1.0 trans-(±)-ACPD A-155 35 1.0 dihydrochloride O 5889 35 1.0 N6-Cyclopentyladenosine C 8031 35 1.0 Cantharidic Acid C 8088 35 1.0 Sodium&Taurocholate&hydrate T&9034 35 1.0 Nisoxetine hydrochloride N-151 36 0.9 hydrochloride M 9651 36 0.9 1-(5-Isoquinolinylsulfonyl)-3-methylpiperazine dihydrochloride I 6391 36 0.9 H-8 dihydrochloride M 9656 36 0.9 Naloxone benzoylhydrazone N-165 36 0.9 NS-1619 N-170 36 0.9 dihydrochloride N-176 36 0.9 NBQX disodium N-183 36 1.0 (-)- sodium M 1275 36 1.0 2-Methyl-5-hydroxytryptamine maleate M-109 36 0.9 Metolazone M-116 36 1.0

18 Table&S2

(Cy53dT153NS3h(Binding( (Cy53dT153NS3h(Binding( Compound(Name( Sigma(Cat.(No. %(Inhibition Interference( N-omega-Methyl-5-hydroxytryptamine oxalate salt M 1514 36 0.9 hydrochloride O 2378 36 0.9 S(+)-Ibuprofen I-106 36 0.9 p-Iodoclonidine hydrochloride I-114 36 0.9 M3071 36 0.9 R(+)-IAA-94 I-117 36 1.0 O 8757 36 0.9 Piceatannol P 0453 36 0.9 TBB T 0826 36 1.0 P 0667 36 1.0 CR 2249 N 2034 36 1.0 hydrochloride N 3136 36 1.0 K 3375 36 1.0 Clorgyline hydrochloride M 3778 36 1.0 SKF-525A hydrochloride P 1061 36 0.9 hydrochloride M 5154 36 1.0 Gabapentin G-154 36 1.0 ABT-702 dihydrochloride A2721 36 0.9 Guanidinyl-naltrindole di-trifluoroacetate G 3416 36 0.9 Raloxifene hydrochloride R 1402 36 0.9 mesylate P 7561 36 0.9 S-103 36 0.9 dihydrochloride T 8516 36 0.9 SDZ-205,557 hydrochloride S-174 36 1.0 Ribavirin R 9644 36 0.9 Tropicamide T 9778 36 0.9 methanesulfonate P 8828 36 1.0 FAUC 213 F 4429 36 0.9 (±)-Thalidomide T-144 36 1.0 SR 57227A S 1688 36 1.0 (±)-Chlorpheniramine maleate C 3025 36 0.9 ABT-418 hydrochloride A 6476 36 1.0 hydrochloride B 5683 36 0.9 9-cyclopentyladenine C 4479 36 0.9 (+)-Chlorpheniramine maleate C 4915 36 0.9 hydrochloride hemihydrate A 4393 36 1.0 CHM-1 hydrate C1244 37 0.9 hydrochloride I 6138 37 0.9 hydrochloride I 7379 37 1.0 Linopirdine L-134 37 0.9 alpha-Methyl-5-hydroxytryptamine maleate M-110 37 0.9 Sodium Oxamate O 2751 37 0.9 MDL 26,630 trihydrochloride M-166 37 0.9 Palmitoylethanolamide P 0359 37 1.0 Ranolazine dihydrochloride R 6152 37 0.9 3-Tropanyl-3,5-dichlorobenzoate T 8160 37 0.9 R 6520 37 1.0 L-741,626 L-135 38 0.9 L-703,606 oxalate salt hydrate L119 38 0.9 R(-)-N6-(2-Phenylisopropyl)adenosine P 4532 38 1.0 ( R)-(+)-WIN 55,212-2 mesylate W-102 38 1.0 8-(4-Chlorophenylthio)-cAMP sodium C 3912 38 0.9 Amsacrine hydrochloride A 9809 38 0.9 hydrochloride P 6656 39 0.9 AGK2 A 8231 312 0.8 Ruthenium red R 2751 398 0.0

19 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

50930730 3&±&1 201&±&119

46839370 5&±&1 179&±&239

50930756 22&±&2 173&±&354

50930737 4&±&1 128&±&57

50930751 9&±&3 71&±&13

53377551 81&±&22 57&±&13

53239937 4&±&0 57&±&16

50930734 52&±&20 55&±&20

49849302 3&±&0 46&±&52

1 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

50930749 22&±&4 36&±&7

49849299 8&±&1 32&±&7

49849293 4&±&0 28&±&9

50930740 5&±&4 27&±&6

53377547 12&±&4 27&±&5

53377546 29&±&7 23&±&5

53308658 10&±&2 23&±&6

49849293 3&±&1 22&±&4

53377548 44&±&12 20&±&4

2 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

44251433 122&±&5 19&±&8

53377553 29&±&4 17&±&5

53255450 227&±&107 17&±&4

53255447 5&±&3 15&±&3

50930733 19&±&15 14&±&2

49849294 5&±&1 14&±&4

50930745 17&±&6 13&±&1

50930748 14&±&1 13&±&2

44251427 70&±&31 13&±&4

3 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

50930741 6&±&2 12&±&8

53356770 8&±&2 11&±&2

49849298 5&±&1 10&±&3

50930743 17&±&17 10&±&1

49849295 24&±&2 10&±&2

53308659 5&±&0 9&±&2

52914816 14&±&0 9&±&2

44251428 95&±&14 9&±&1

53356771 9&±&4 7&±&1

4 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

53377543 3&±&0 7&±&3

44251429 33&±&24 7&±&2

53356772 4&±&1 6&±&1

49849287 4&±&2 5&±&1

53312458 11&±&2 5&±&1

415713 12&±&1 5&±&1

53356653 4&±&1 5&±&1

49849286 3&±&0 5&±&2

49849284 11&±&7 4&±&1

5 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

Primulin 10&±&5 4&±&1

49849280 11&±&2 4&±&1

53356656 5&±&2 4&±&1

49849276 2&±&0 4&±&2

49849289 6&±&2 3&±&0

49849300 10&±&2 3&±&1

49849282 10&±&3 2&±&1

44251428 6&±&2 2&±&1

44251431 5&±&2 2&±&0

6 Table&S3

DNA*Unwinding* *Cy5;dT15;SSB*Binding* Structure CID*No. IC50,*µM IC50,*µM

49849290 10&±&5 2&±&0

44251434 7&±&4 1&±&0

7