Structural and functional differentiation of each subunit in dimeric HIV-1 protease by single-chain derivatization

Adachi, Motoyasu; Shibazaki, Chie; Arai, Shigeki; Shimizu, Rumi; Kuroki, Ryota

HIV protease-I (HIV-PR) is an important drug target protein for AIDS. Since HIV-PR has a homo dimeric structure, the contribution of each monomer on its structure and function is indistinguishable. To solve this problem, a single chain derivative of HIV-PR (sc-HIV-PR) was designed. The gene coding sc-HIV-PR, in which a linker comprising glycyl-analine was placed between the C-terminal and N-terminal of each HIV-PR, was expressed in E. coli. The inactive form in inclusion bodies was successfully refolded to its active form as reported previously. The tertiary structure of sc-HIV-PR with KNI272 was determined to 1.3 resolution by X-ray crystallography. The structure of scHIV-PR was confirmed to be essentially equivalent to the original dimer form of HIV-PR with a RMSD of less than 0.3 including the structure of active site. This result indicates that the single chain derivatization of HIV-PR was successful and the inserted linker did not affect the overall structure of HIV-PR.