Composition of NS3-4A and the NS3-4A expression construct. (A and B) The NS3-4A complex organization and construct design is illustrated schematically. In panel A, “pro” refers to the serine protease domain, and the Roman numerals indicate the respective NS3 helicase subdomains. The regions where ATP, RNA, and the NS4A cofactor bind are indicated as well. The protein construct expressed in E. coli is depicted in panel B. The numbers below the map refer to the HCV polyprotein numbering of the amino acids of NS3-4A. A detailed map of NS4A (i.e., HCV polyprotein residues 1658 to 1711) (18) is shown below the His-SUMO-NS3-4A expression construct. Residues Y1702 and M1708 are indicated in white font.