Insights into the effect of detergents on the full-length rhomboid protease from Pseudomonas aeruginosa and its cytosolic domain

Sherratt, Allison R.; Braganza, Michael V.; Nguyen, Elizabeth; Ducat, Thierry; Goto, Natalie K.

doi:10.1016/j.bbamem.2009.09.003

Cited by 19 publications

(20 citation statements)

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“…These domains are often large and conserved between rhomboid family members but are not essential for catalysis, suggesting a function in substrate recognition/specificity or regulation of activity (reviewed in [23]). Consistent with this hypothesis, the removal of the Nterminal cytosolic domain of RHBDL2 affects its activity in vitro [19] and the N-terminal domain of the Pseudomonas aeruginosa GlpG is also important for its function in vitro [25]. In the case of GlpG, Sherratt et al suggested that the N-terminus of the protease interacts with the catalytic core domain, enhancing its activity by altering the conformation/stability of the active site and/or aiding anchorage of the protease to the membrane [25].…”

Section: Cleavage Site and Substrate Recognitionsupporting

confidence: 60%

“…Consistent with this hypothesis, the removal of the Nterminal cytosolic domain of RHBDL2 affects its activity in vitro [19] and the N-terminal domain of the Pseudomonas aeruginosa GlpG is also important for its function in vitro [25]. In the case of GlpG, Sherratt et al suggested that the N-terminus of the protease interacts with the catalytic core domain, enhancing its activity by altering the conformation/stability of the active site and/or aiding anchorage of the protease to the membrane [25]. The iRhom family of pseudoproteases, which evolved from rhomboid proteases that lost their catalytic activity but retained their localization and the ability to bind their substrates, may provide additional regulatory functions.…”

Section: Cleavage Site and Substrate Recognitionsupporting

confidence: 60%

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New insights into parasite rhomboid proteases

Santos

Graindorge

Soldati‐Favre

2012

Molecular and Biochemical Parasitology

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a b s t r a c tThe rhomboid-like proteins constitute a large family of intramembrane serine proteases that are present in all branches of life. First studied in Drosophila, these enzymes catalyse the release of the active forms of proteins from the membrane and hence trigger signalling events. In protozoan parasites, a limited number of rhomboid-like proteases have been investigated and some of them are associated to pathogenesis. In Apicomplexans, rhomboid-like protease activity is involved in shedding adhesins from the surface of the zoites during motility and host cell entry. Recently, a Toxoplasma gondii rhomboid was also implicated in an intracellular signalling mechanism leading to parasite proliferation. In Entamoeba histolytica, the capacity to adhere to host cells and to phagocytose cells is potentiated by a rhomboid-like protease. Survey of a small number of protozoan parasite genomes has uncovered species-specific rhomboidlike protease genes, many of which are predicted to encode inactive enzymes. Functional investigation of the rhomboid-like proteases in other protozoan parasites will likely uncover novel and unexpected implications for this family of proteases.

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Section: Cleavage Site and Substrate Recognitionsupporting

confidence: 60%

Section: Cleavage Site and Substrate Recognitionsupporting

confidence: 60%

New insights into parasite rhomboid proteases

Santos

Graindorge

Soldati‐Favre

2012

Molecular and Biochemical Parasitology

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“…has been reported (21,35). Here, we see that the alkyl chain length does not have a significant effect on the rhomboid activity, unless the chains are very short compared with the natural E. coli phospholipids.…”

Section: Compoundsupporting

confidence: 50%

Activity-based probes for rhomboid proteases discovered in a mass spectrometry-based assay

Vosyka

Vinothkumar

Wolf

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

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Rhomboid proteases are evolutionary conserved intramembrane serine proteases. Because of their emerging role in many important biological pathways, rhomboids are potential drug targets. Unfortunately, few chemical tools are available for their study. Here, we describe a mass spectrometry-based assay to measure rhomboid substrate cleavage and inhibition. We have identified isocoumarin inhibitors and developed activity-based probes for rhomboid proteases. The probes can distinguish between active and inactive rhomboids due to covalent, reversible binding of the active-site serine and stable modification of a histidine residue. Finally, the structure of an isocoumarin-based inhibitor with Escherichia coli rhomboid GlpG uncovers an unusual mode of binding at the active site and suggests that the interactions between the 3-substituent on the isocoumarin inhibitor and hydrophobic residues on the protease reflect S′ subsite binding. Overall, these probes represent valuable tools for rhomboid study, and the structural insights may facilitate future inhibitor design.MALDI screening | covalent inhibition | regulated intramembrane proteolysis P roteolysis controls many important biological processes, such as apoptosis, antigen presentation, and blood coagulation. Selective digestion of protein substrates is possible by a combination of tight posttranslational control of protease activity (1) and the protease's substrate specificity, which generally is governed by the primary sequence around the scissile bond (2). The use of inhibitors and activity-based probes (ABPs) has led to a tremendous gain in understanding the roles of proteases within physiological and pathological processes (3). ABPs are small molecules that bind only to active enzymes, but not to zymogen or inhibitor-bound forms (4). ABPs generally consist of a detection tag, a spacer, and a "warhead." The warhead covalently binds to the target enzyme(s) and often is derived from a mechanism-based inhibitor. In the past, ABPs were used to study the activation, localization, and function of soluble proteases in a variety of organisms and disease models (5).Most proteases are soluble and surrounded by an aqueous environment. However, several families of intramembrane proteases exist (6-8): the metalloprotease family M50 (site-2 protease), the aspartic protease family A22 (signal peptide peptidase and γ-secretase), and the serine protease family S54 [rhomboid; numbering according to the MEROPS database (9)]. Rhomboid was discovered in 2001 as a protease in the EGF receptor signaling pathway in the fruitfly Drosophila melanogaster (10). Interestingly, rhomboid genes occur in all kingdoms of nature and are found in most sequenced organisms (11,12). Rhomboids appear to have a wide range of physiological functions, including bacterial protein export (13) and invasion by apicomplexan parasites (14,15), but the roles of many rhomboids remain to be discovered.Rhomboids catalyze peptide bond hydrolysis using a catalytic dyad formed by a serine residue in transmembrane domain...

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“…The rhomboid protein is an ancient conserved family of intramembrane serine proteases that catalyze the cleavage of transmembrane segments within the lipid membrane to achieve a wide range of biological functions (Sherrat et al, 2009). Recent reports have indicated that a subfamily of rhomboids are gatekeepers of mitochondrial dynamics and apoptosis, thus introducing a new paradigm of how the mitochondria uses these proteases to direct the stress responses, signal to the nucleus, and other key mitochondrial activities in health and disease (Hill and Pellegrini, 2010).…”

Section: Organelle-related Transcriptsmentioning

confidence: 99%