Structure, interdomain dynamics and pH-dependent autoactivation of pro-rhodesain, the main lysosomal cysteine protease from African trypanosomes

  • Rhodesain is the lysosomal cathepsin L-like cysteine protease of T. brucei rhodesiense, the causative agent of Human African Trypanosomiasis. The enzyme is essential for the proliferation and pathogenicity of the parasite as well as its ability to overcome the blood-brain barrier of the host. Lysosomal cathepsins are expressed as zymogens with an inactivating pro-domain that is cleaved under acidic conditions. A structure of the uncleaved maturation intermediate from a trypanosomal cathepsin L-like protease is currently not available. We thus established the heterologous expression of T. brucei rhodesiense pro-rhodesain in E. coli and determined its crystal structure. The trypanosomal pro-domain differs from non-parasitic pro-cathepsins by a unique, extended α-helix that blocks the active site and whose interactions resemble that of the antiprotozoal inhibitor K11777. Interdomain dynamics between pro- and core protease domain as observed by photoinduced electron transfer fluorescence correlation spectroscopy increase at low pH, where pro-rhodesain also undergoes autocleavage. Using the crystal structure, molecular dynamics simulations and mutagenesis, we identify a conserved interdomain salt bridge that prevents premature intramolecular cleavage at higher pH values and may thus present a control switch for the observed pH-sensitivity of pro-enzyme cleavage in (trypanosomal) CathL-like proteases.

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Author:Patrick JohéGND, Elmar JaenickeORCiDGND, Hannes NeuweilerORCiDGND, Tanja SchirmeisterORCiDGND, Christian KerstenORCiDGND, Ute HellmichORCiDGND
Parent Title (English):bioRxiv
Document Type:Preprint
Date of Publication (online):2020/11/10
Date of first Publication:2020/11/10
Publishing Institution:Universitätsbibliothek Johann Christian Senckenberg
Release Date:2023/07/04
Tag:African Sleeping Sickness; Trypanosoma brucei; autoinhibition; pro-domain; pro-enzyme; rhodesain; zymogen
Page Number:29
Institutes:Wissenschaftliche Zentren und koordinierte Programme / Zentrum für Biomolekulare Magnetische Resonanz (BMRZ)
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Licence (German):License LogoCreative Commons - CC BY-NC-ND - Namensnennung - Nicht kommerziell - Keine Bearbeitungen 4.0 International