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. 2015 Aug;71(Pt 8):1055-62.
doi: 10.1107/S2053230X15011127. Epub 2015 Jul 29.

Structure of BbKI, a disulfide-free plasma kallikrein inhibitor

Dongwen Zhou  1 , Daiane Hansen  2 , Ivan G Shabalin  3 , Alla Gustchina  1 , Debora F Vieira  4 , Marlon V de Brito  4 , Ana Paula U Araújo  4 , Maria Luiza V Oliva  2 , Alexander Wlodawer  1
Affiliations

Structure of BbKI, a disulfide-free plasma kallikrein inhibitor

Dongwen Zhou et al. Acta Crystallogr F Struct Biol Commun. 2015 Aug.

Abstract

A serine protease inhibitor from Bauhinia bauhinioides (BbKI) belongs to the Kunitz family of plant inhibitors, which are common in plant seeds. BbKI does not contain any disulfides, unlike most other members of this family. It is a potent inhibitor of plasma kallikrein, in addition to other serine proteases, and thus exhibits antithrombotic activity. A high-resolution crystal structure of recombinantly expressed BbKI was determined (at 1.4 Å resolution) and was compared with the structures of other members of the family. Modeling of a complex of BbKI with plasma kallikrein indicates that changes in the local structure of the reactive loop that includes the specificity-determining Arg64 are necessary in order to explain the tight binding. An R64A mutant of BbKI was found to be a weaker inhibitor of plasma kallikrein, but was much more potent against plasmin, suggesting that this mutant may be useful for preventing the breakup of fibrin and maintaining clot stability, thus preventing excessive bleeding.

Keywords: Kunitz inhibitor; crystal structure; kallikrein; β-trefoil.

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Figures

Figure 1
Figure 1
A stereoimage showing a fragment of the final model covered by the 2F o − F c electron-density map, contoured at 1σ, in the part of the reactive loop that includes the specificity-determining Arg64. Two orientations of the main chain of this residue are clearly visible.
Figure 2
Figure 2
A stereoimage of BbKI viewed along the pseudo-threefold axis. The three β-sheets forming the core of a β-trefoil are colored blue, purple and green. The reactive loop responsible for the inhibitory properties of the inhibitor is red, and the specificity-determining Arg64 is shown in stick representation.
Figure 3
Figure 3
A stereoimage showing the superposition of the main chains of BbKI (grey) and BbCI (green) in an orientation similar to that in Fig. 2 ▸. The reactive loop of BbKI responsible for the inhibitory properties is shown in red; the specificity-determining Arg64 in BbKI is shown in stick representation. The four loops in BbKI that differ most from BbCI (see text for details) are shown in blue and labeled.
Figure 4
Figure 4
A stereoimage showing the hydrophobic environment of the sole cysteine in BbKI (Cys155).
See this image and copyright information in PMC

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