title: Conformational changes of rBTI from buckwheat upon binding to trypsin: implications for the role of the P(8)' residue in the potato inhibitor I family.
abstract: BWI-1 (buckwheat trypsin inhibitor), a member of the potato inhibitor I family, suppresses the growth of T-acute lymphoblastic leukemia cells and induces apoptosis in human solid tumor cell lines. Here, we report the crystal structure of rBTI (recombinant buckwheat trypsin inhibitor), a recombinant protein of BWI-1, at 1.84 Å resolution and the structure of rBTI in complex with bovine trypsin at 2.26 Å resolution. A conformational change of Trp53 at the P(8)' position in rBTI was observed upon its binding to trypsin, which is not seen in other members of the potato inhibitor I family reported previously. The role of the P(8)' residue in the potato inhibitor I family was examined by measuring the association and dissociation rates of four rBTI mutants with different substitutions at the P(2) and P(8)' positions when binding to trypsin. One of the mutants, P44T, was found to be a much stronger inhibitor than wild-type rBTI, with a picomolar (pM) dissociation constant. Our results could provide valuable insights for designing a new rBTI-based antitumor drug in the future.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing, People's Republic of China.
authors.forename: Longfei
authors.initials: L
authors.lastname: Wang
authors.forename: Fei
authors.initials: F
authors.lastname: Zhao
authors.forename: Mei
authors.initials: M
authors.lastname: Li
authors.forename: Hongmei
authors.initials: H
authors.lastname: Zhang
authors.forename: Yu
authors.initials: Y
authors.lastname: Gao
authors.forename: Peng
authors.initials: P
authors.lastname: Cao
authors.forename: Xiaowei
authors.initials: X
authors.lastname: Pan
authors.forename: Zhuanhua
authors.initials: Z
authors.lastname: Wang
authors.forename: Wenrui
authors.initials: W
authors.lastname: Chang
mesh_terms: D000818:Animals; D002417:Cattle; D006860:Hydrogen Bonding; D008958:Models, Molecular; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014357:Trypsin / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000737:chemistry* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D014361:Trypsin Inhibitors; D014357:Trypsin