Unbound form of tomato inhibitor-II reveals interdomain flexibility and conformational variability in the reactive site loops.
The Potato II (Pot II) family of proteinase inhibitors plays important roles in the constitutive and inducible defense of plants against predation by a wide range of pests. The structural basis of inhibition by a multidomain Pot II family inhibitor was revealed recently by the structure of the ternary complex between the two-headed tomato inhibitor-II (TI-II) and two molecules of subtilisin Carlsberg. Here we report the 2.15-A resolution crystal structure of the unbound form of TI-II that reveals significant conformational flexibility in the absence of bound proteinase molecules. The four independent copies of unbound TI-II in the asymmetric unit of the unit cell display a range of different conformations when compared with the bound form of the inhibitor, most strikingly in the orientations of the inhibitory domains and in the conformations of the reactive site loops. One of the two linker segments (residues 74 to 79) between the two domains as well as the adjacent beta-strand in Domain I (residues 80-85) is well ordered in all four copies of the unbound inhibitor, even though this region appeared to be disordered in the structure of the ternary complex. Conformational flexibility seen in the reactive site loops of unbound TI-II suggests a mechanism by which the inhibitor can balance the need for tight binding with the need for broad inhibitory function.
Barrette-Ng
Isabelle H
IH
Canadian Institutes of Health Research Group in Protein Structure and Function, Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Ng
Kenneth K-S
KK
Cherney
Maia M
MM
Pearce
Gregory
G
Ghani
Usman
U
Ryan
Clarence A
CA
James
Michael N G
MN
D001665:Binding Sites; D018360:Crystallography, X-Ray; D018551:Solanum lycopersicum / Q000201:enzymology* / Q000235:genetics; D009097:Multienzyme Complexes / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011480:Protease Inhibitors / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary
D009097:Multienzyme Complexes; D010940:Plant Proteins; D011480:Protease Inhibitors; C073705:prosystemin protein, Lycopersicon esculentum