Supporting information for Disulfiram can inhibit MERS and SARS coronavirus papain-like proteases via different modes
Min-Han Lin1, David C. Moses2, Chih-Hua Hsieh1, Shu-Chun Cheng3, Yau-Hung Chen2, Chiao-Yin Sun3,*, Chi-Yuan Chou1,*
1 Department of Life Sciences and Institute of Genome Sciences, National YangMing University, Taipei 112, Taiwan 2 Department of Chemistry, Tamkang University, Tamsui, Taiwan 3 Department of Nephrology, Chang-Gung Memorial Hospital, Keelung 204, Taiwan
_____________________ *Correspondence information for Dr. Chi-Yuan Chou. Address: 155 Li-Nong St., Sec. 2, Taipei 112, Taiwan, R.O.C. Phone: +886-2-28267168. FAX: +886-2-28202449. Email:
[email protected] and Dr. Chiao-Yin Sun. Address: 222 Mai-Chin Rd., Keelung 204, Taiwan, R. O. C. Phone: +886-2-24313131 ext. 3170. FAX: +886-224335342. E-mail:
[email protected].
60
1/vi (min/M)
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Disulfiram 0 M 6 M 24 M 36 M 50 M
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-0.04
-0.02
0.00
0.02
0.04
0.06
0.08
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1/[Peptide substrate] (M-1)
Figure S1. Inhibitory effect of SARS-CoV PLpro by disulfiram was fitted to a mixed inhibition model. The proteolytic activity of the enzyme was measured in the presence of 10-50 μM peptide substrate and 6-50 μM disulfiram, respectively. The solid line are the best-fit results in accordance with a mixed inhibition model (Eq. 4). The Rsqr value is 0.979. Kinetic parameters from the best-fit results are shown in Table 1.
A
C
B
D
Figure S2. Active site and zinc-binding site of SARS-CoV PLpro in complex with βME (A and C) and glycerol (B and D). (A) The 2Fo-Fc electron density at 1.65 Å resolution of βME and the sulfur of Cys112 contoured at 1.0σ. (B) The omit Fo-Fc electron density at 1.65 Å of glycerol contoured at 3.0σ. The red dashed lines indicate hydrophilic interactions. (C) and (D) The 2Fo-Fc electron density at 1.65 Å resolution of zinc ion and the four coordinated cysteines contoured at 1.0σ.
SARS-CoV PLpro C271A mutant
vi (Intensity/s)
0.12 0.10
IC50: 62.7 ± 2.0 μM
0.08 0.06 0.04 0.02 0.00 0
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[Disulfiram] (M) Figure S3. Inhibitory effect of disulfiram on SARS-CoV PLpro C271A mutant. Proteolytic activity of SARS-CoV PLpro C271A was measured in the absence or presence of various concentrations of disulfiram (0-200 μM). The concentrations of peptidyl substrate and enzyme were 25 μM and 0.05 μM, respectively. The line shows the best-fit result in accordance with the IC50 equation.
