Table 1. Mutants used in this study.

		Mutations a
Gene	Name	JFH-1	H77	Rationale	References
NS5B	5Bm1	D318N	D2738	NS5B polymerase active site, Mg++ coordination	[25,34,67]
		D319N	D2739
	5Bm2	R168A	R2588	NS5B RNA binding groove	[68,69]
		K172A	K2592
	5Bm3	As above		Combined 5Bm1 and 5Bm2	This study
	5A*5B	S1	S2421	Nonsense mutation at NS5B start
	5Brc	n.a.		Recoded NS5B to create -1 ORF	This study
	5Bfs	T40–R592		-1 Frameshift at NS5B codon 40	This study
NS3	3m1	H57A	H1083	Serine protease active site	[70]
	3m2	S139A	S1165	Serine protease active site	[70,71]
	3m3	T269A	T1295	Helicase RNA binding	[72,73]
	3m4	D290A	D1316	Helicase NTPase active site	[34,72,74]
		E291A	D1317
	3m5	W501A	W1527	Helicase stacking	[73,75–77]
	3m6	ΔF184	F1210	Linker between the NS3 serine protease and helicase domains	[78]
		ΔS185	T1211
		ΔD186	D1212
	3m7	P190G	P1216	Linker between the NS3 serine protease and helicase domains	[78]
		P191G	P1217
NS4A	4Am1	Y45A	Y1702	Essential, conserved Tyr in NS4A C-terminal acidic domain	[65,79]
	4Am2	A11L	A1668	NS4A transmembrane dimerization	[80]
NS4B	4Bm1	E226A	E1937	Determinant of NS4B homotypic interaction and membrane alteration	[40,58]
NS5A	5Am1	S232I	S2204	Determinant of NS5A hyperphosphorylation	[58,81,82]
	DCVR	F28S	M2000	Daclatasvir resistance	[59]
NS3–NS5A	3–5Arc	n.a.		Recoded NS3 codon 111 to NS5A codon 120	This study
NS3, NS5B	3&5Bm	As above		Combined 3m6 and 5Bm1	This study
NS4A–NS5A	4A–5Am	As above		Combined 4Am1, 4Bm1, and 5Am1	This study
NS3–NS5B	3–5Bm	As above		Combined 3m6, 4Am1, 4Bm1, 5Am1, and 5Bm1	This study

^aResidues are numbered by their codon position within each gene of the JFH-1 subgenomic replicon or in the polyprotein of HCV reference strain H77; n.a., not applicable.