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A Guide to Production, Crystallization, and Structure Determination of Human IKK1/α
Published on: November 2, 2018
Direct and indirect mechanisms of KLK4 inhibition revealed by structure and dynamics
Blake T Riley1, Olga Ilyichova1, Mauricio G S Costa2
1Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Clayton, Victoria 3800, Australia.
Abstract:
The kallikrein-related peptidase (KLK) family of proteases is involved in many aspects of human health and disease. One member of this family, KLK4, has been implicated in cancer development and metastasis. Understanding mechanisms of inactivation are critical to developing selective KLK4 inhibitors. We have determined the X-ray crystal structures of KLK4 in complex with both sunflower trypsin inhibitor-1 (SFTI-1) and a rationally designed SFTI-1 derivative to atomic (~1 Å) resolution, as well as with bound nickel. These structures offer a structural rationalization for the potency and selectivity of these inhibitors, and together with MD simulation and computational analysis, reveal a dynamic pathway between the metal binding exosite and the active site, providing key details of a previously proposed allosteric mode of inhibition. Collectively, this work provides insight into both direct and indirect mechanisms of inhibition for KLK4 that have broad implications for the enzymology of the serine protease superfamily, and may potentially be exploited for the design of therapeutic inhibitors.
Insights
Researchers elucidated the inactivation mechanisms of kallikrein-related peptidase 4 (KLK4) using X-ray crystallography. This study reveals how inhibitors bind to KLK4, aiding in the development of targeted cancer therapies.
Area of Science:
- Biochemistry and Structural Biology
- Enzymology
- Protease Inhibitor Design
Background:
- The kallikrein-related peptidase (KLK) family, including KLK4, plays roles in human health and disease.
- KLK4 is implicated in cancer development and metastasis.
- Understanding KLK4 inactivation is crucial for developing selective inhibitors.
Purpose of the Study:
- To determine the X-ray crystal structures of KLK4 in complex with inhibitors.
- To elucidate the mechanisms of KLK4 inhibition.
- To provide a structural basis for designing KLK4 inhibitors.
Main Methods:
- X-ray crystallography of KLK4 complexed with sunflower trypsin inhibitor-1 (SFTI-1) and a derivative.
- Determination of structures to atomic resolution (~1 Å).
- Molecular dynamics (MD) simulations and computational analysis.
Main Results:
- Atomic resolution structures of KLK4 with SFTI-1 and a derivative were determined.
- Structural rationalization for inhibitor potency and selectivity was provided.
- A dynamic pathway between the metal binding exosite and active site was revealed, detailing an allosteric inhibition mode.
Conclusions:
- The study provides insight into direct and indirect KLK4 inhibition mechanisms.
- Findings have broad implications for serine protease superfamily enzymology.
- The results can be exploited for the design of therapeutic KLK4 inhibitors.
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