Solution structure of a phytocystatin from Ananas comosus and its molecular interaction with papain.

The structure of a recombinant pineapple cystatin (AcCYS) was determined by NMR with the RMSD of backbone and heavy atoms of twenty lowest energy structures of 0.56 and 1.11 Å, respectively. It reveals an unstructured N-terminal extension and a compact inhibitory domain comprising a four-stranded antiparallel β-sheet wrapped around a central α-helix. The three structural motifs (G(45), Q(89)XVXG, and W(120)) putatively responsible for the interaction with papain-like proteases are located in one side of AcCYS. Significant chemical shift perturbations in two loop regions, residues 45 to 48 (GIYD) and residues 89 to 91 (QVV), of AcCYS strongly suggest their involvement in the binding to papain, consistent with studies on other members of the cystatin family. However, the highly conserved W120 appears not to be involved in the binding with papain as no chemical shift perturbation was observed. Chemical shift index analysis further indicates that the length of the α-helix is shortened upon association with papain. Collectively, our data suggest that AcCYS undergoes local secondary structural rearrangements when papain is brought into close contact. A molecular model of AcCYS/papain complex is proposed to illustrate the interaction between AcCYS and papain, indicating a complete blockade of the catalytic triad by AcCYS.

Abstract licence: CC BY

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