Kvido Stříšovský, PhD
Substrate binding and specificity of rhomboid intramembrane protease revealed by substrate-peptide complex structures
EMBO Journal 33 (20): 2408-2421 (2014).
The mechanisms of intramembrane proteases are incompletely understood due to the lack of structural data on substrate complexes. To gain insight into substrate binding by rhomboid proteases, we have synthesised a series of novel peptidyl‐chloromethylketone (CMK) inhibitors and analysed their interactions with Escherichia coli rhomboid GlpG enzymologically and structurally. We show that peptidyl‐CMKs derived from the natural rhomboid substrate TatA from bacterium Providencia stuartii bind GlpG in a substrate‐like manner, and their co‐crystal structures with GlpG reveal the S1 to S4 subsites of the protease. The S1 subsite is prominent and merges into the ‘water retention site’, suggesting intimate interplay between substrate binding, specificity and catalysis. Unexpectedly, the S4 subsite is plastically formed by residues of the L1 loop, an important but hitherto enigmatic feature of the rhomboid fold. We propose that the homologous region of members of the wider rhomboid‐like protein superfamily may have similar substrate or client‐protein binding function. Finally, using molecular dynamics, we generate a model of the Michaelis complex of the substrate bound in the active site of GlpG.
The Rhomboid Superfamily: Structural Mechanisms and Chemical Biology Opportunities
Trends in Biochemical Sciences 43 (9): 726-739 (2018).
General and Modular Strategy for Designing Potent, Selective, and Pharmacologically Compliant Inhibitors of Rhomboid Proteases
Cell Chemical Biology 24 (12): 1523-1536 (2017).
Quantitative proteomics screen identifies a substrate repertoire of rhomboid protease RHBDL2 in human cells and implicates it in epithelial homeostasis
Scientific Reports 7 : 7283 (2017).
Sensitive Versatile Fluorogenic Transmembrane Peptide Substrates for Rhomboid Intramembrane Proteases
Journal of Biological Chemistry 292 (7): 2703-2713 (2017).