Activation Route of the Schistosoma mansoni Cathepsin B1 Drug Target: Structural Map with a Glycosaminoglycan Switch

Jílková, Adéla; Horn, Martin; Řezáčová, P.; Marešová, Lucie; Fajtová, Pavla; Brynda, J.; Vondrášek, Jiřı́; McKerrow, James H.; Caffrey, Conor R.

doi:10.1016/j.str.2014.09.015

Cited by 33 publications

(39 citation statements)

References 46 publications

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“…As this motif occurs in all TrCB1 prosequences (Figure 2), sulfated polysaccharides facilitated auto-activation of both TrCB1.1 and TrCB1.4 (Figure 1). However, unlike in SmCB1 (Jílková et al, 2014), the products were relatively unstable at all tested pH values. Therefore, we did not use them in most subsequent experiments.…”

Section: Discussionmentioning

confidence: 71%

“…Auto-activation of recombinant cathepsin B1 from Schistosoma mansoni (SmCB1) is forced by negatively charged polysaccharides, such as sulfated glycosaminoglycans. Their binding to the SmCB1 pro-sequence destabilizes the pro-sequence/enzyme core junction (Jílková et al, 2014) and is mediated by a specific α-helix that contains a "heparin-binding motif "-XBBXBBX-(B is a basic amino acid and X refers to a hydropathic residue) (Horn et al, 2011;Jílková et al, 2014). A sequence alignment revealed that this motif is specific for trematode cathepsins B involved in the digestion of host proteins (Horn et al, 2011).…”

Section: Discussionmentioning

confidence: 99%

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Isoforms of Cathepsin B1 in Neurotropic Schistosomula of Trichobilharzia regenti Differ in Substrate Preferences and a Highly Expressed Catalytically Inactive Paralog Binds Cystatin

Dvořáková

Leontovyč

Macháček

et al. 2020

Front. Cell. Infect. Microbiol.

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Schistosomula (the post-infective stages) of the neurotropic schistosome Trichobilharzia regenti possess multiple isoforms of cathepsin B1 peptidase (TrCB1.1-TrCB1.6) with involvement in nutrient digestion. The comparison of substrate preferences of TrCB1.1 and TrCB1.4 showed that TrCB1.4 had a very narrow substrate specificity and after processing it was less effective toward protein substrates when compared to TrCB1.1. Self-processing of both isoforms could be facilitated by sulfated polysaccharides due to a specific binding motif in the pro-sequence. Trans-activation by heterologous enzymes was also successfully employed. Expression profiling revealed a high level of transcription of genes encoding the enzymatically inactive paralogs TrCB1.5 and TrCB1.6. The transcription level of TrCB1.6 was comparable with that of TrCB1.1 and TrCB1.2, the most abundant active isoforms. Recombinant TrCB1.6wt, a wild type paralog with a Cys 29-to-Gly substitution in the active site that renders the enzyme inactive, was processed by the active TrCB1 forms and by an asparaginyl endopeptidase. Although TrCB1.6wt lacked hydrolytic activity, endopeptidase, but not dipeptidase, activity could be restored by mutating Gly 29 to Cys 29. The lack of exopeptidase activity may be due to other mutations, such as His 110-to-Asn in the occluding loop and Asp 224-to-Gly in the main body of the mature TrCB1.6, which do not occur in the active isoforms TrCB1.1 and TrCB1.4 with exopeptidase activity. The catalytically active enzymes and the inactive TrCB1.6 paralog formed complexes with chicken cystatin, thus supporting experimentally the hypothesis that inactive paralogs could potentially regulate the activity of the active forms or protect them from being inhibited by host inhibitors. The effect on cell viability and nitric oxide production by selected immune cells observed for TrCB1.1 was not confirmed for TrCB1.6. We show here that the active isoforms of TrCB1 have different affinities for peptide substrates thereby facilitating diversity in protein-derived nutrition for Dvořáková et al. Trichobilharzia Cathepsin B1 Paralogs the parasite. The inactive paralogs are unexpectedly highly expressed and one of them retains the ability to bind cystatins, likely due to specific mutations in the occluding loop and the enzyme body. This suggests a role in sequestration of inhibitors and protection of active cysteine peptidases.

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Section: Discussionmentioning

confidence: 71%

Section: Discussionmentioning

confidence: 99%

Isoforms of Cathepsin B1 in Neurotropic Schistosomula of Trichobilharzia regenti Differ in Substrate Preferences and a Highly Expressed Catalytically Inactive Paralog Binds Cystatin

Dvořáková

Leontovyč

Macháček

et al. 2020

Front. Cell. Infect. Microbiol.

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“…The resulting fragments were purified by RP-HPLC over a 5 mm C18 column (Luna, Phenomenex) equilibrated in 0.1% TFA and eluted with a linear gradient of 90% acetonitrile in 0.1% TFA. The purified peptides were characterized by ESI mass spectrometry on a Q-Tof Micro (Waters) (Jilkova et al, 2014).…”

Section: Declaration Of Interestsmentioning

confidence: 99%

Novel Structural Mechanism of Allosteric Regulation of Aspartic Peptidases via an Evolutionarily Conserved Exosite

Hánová

Brynda

Houštecká

et al. 2018

Cell Chemical Biology

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Pepsin-family aspartic peptidases are biosynthesized as inactive zymogens in which the propeptide blocks the active site until its proteolytic removal upon enzyme activation. Here, we describe a novel dual regulatory function for the propeptide using a set of crystal structures of the parasite cathepsin D IrCD1. In the IrCD1 zymogen, intramolecular autoinhibition by the intact propeptide is mediated by an evolutionarily conserved exosite on the enzyme core. After activation, the mature enzyme employs the same exosite to rebind a small fragment derived from the cleaved propeptide. This fragment functions as an effective natural inhibitor of mature IrCD1 that operates in a pH-dependent manner through a unique allosteric inhibition mechanism. The study uncovers the propeptide-binding exosite as a target for the regulation of pepsin-family aspartic peptidases and defines the structural requirements for exosite inhibition.

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“…The S. mansoni cathepsin B1 (SmCB1), which is a highly abundant digestive protease, was the focal point of various studies (reviewed in Ref. [104]). SmCB1 has been validated as a molecular target for therapy against schistosomiasis in a murine model of S. mansoni infection [105].…”

Section: Peptidasesmentioning

confidence: 99%

Schistosomiasis: Setting Routes for Drug Discovery

Tavares¹,

Gava²,

Oliveira³

et al. 2016

Special Topics in Drug Discovery

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Schistosomiasis is the second most prevalent parasitic disease in the world. Currently, the treatment of this disease relies on a single drug, praziquantel, and due to the identification of resistant parasites, the development of new drugs is urged. The demand for the development of robust high-throughput parasite screening techniques is increasing as drug discovery research in schistosomiasis gains significance. Here, we review the most common methods used for compound screening in the parasites life stages and also summarize some of the methods that have been recently developed. In addition, we reviewed the methods most commonly implemented to search for promising targets and how they have been used to validate new targets against the parasite Schistosoma mansoni. We also review some promising targets in this parasite and show the main approaches and the major advances that have been achieved by those studies. Moreover, we share our experiences in schistosomiasis drug discovery attained with our S. mansoni drug screening platform establishment.

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Activation Route of the Schistosoma mansoni Cathepsin B1 Drug Target: Structural Map with a Glycosaminoglycan Switch

Cited by 33 publications

References 46 publications

Isoforms of Cathepsin B1 in Neurotropic Schistosomula of Trichobilharzia regenti Differ in Substrate Preferences and a Highly Expressed Catalytically Inactive Paralog Binds Cystatin

Isoforms of Cathepsin B1 in Neurotropic Schistosomula of Trichobilharzia regenti Differ in Substrate Preferences and a Highly Expressed Catalytically Inactive Paralog Binds Cystatin

Novel Structural Mechanism of Allosteric Regulation of Aspartic Peptidases via an Evolutionarily Conserved Exosite

Schistosomiasis: Setting Routes for Drug Discovery

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