scholarly journals Human and mouse granzyme M display divergent and species-specific substrate specificities

2011 ◽  
Vol 437 (3) ◽  
pp. 431-442 ◽  
Author(s):  
Stefanie A.H. de Poot ◽  
Marijn Westgeest ◽  
Daniel R. Hostetter ◽  
Petra van Damme ◽  
Kim Plasman ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Specific Substrate ◽  
Structural Determinants ◽  
Cytotoxic Lymphocyte ◽  
Potent Inducer ◽  
Methionine Residue ◽  
Positional Scanning ◽  
Methionine Residues ◽  
Species Specific ◽  
Positional Scanning Libraries

Cytotoxic lymphocyte protease GrM (granzyme M) is a potent inducer of tumour cell death and a key regulator of inflammation. Although hGrM (human GrM) and mGrM (mouse GrM) display extensive sequence homology, the substrate specificity of mGrM remains unknown. In the present study, we show that hGrM and mGrM have diverged during evolution. Positional scanning libraries of tetrapeptide substrates revealed that mGrM is preferred to cleave after a methionine residue, whereas hGrM clearly favours a leucine residue at the P1 position. The kinetic optimal non-prime subsites of both granzymes were also distinct. Gel-based and complementary positional proteomics showed that hGrM and mGrM have a partially overlapping set of natural substrates and a diverged prime and non-prime consensus cleavage motif with leucine and methionine residues being major P1 determinants. Consistent with positional scanning libraries of tetrapeptide substrates, P1 methionine was more frequently used by mGrM as compared with hGrM. Both hGrM and mGrM cleaved α-tubulin with similar kinetics. Strikingly, neither hGrM nor mGrM hydrolysed mouse NPM (nucleophosmin), whereas human NPM was hydrolysed efficiently by GrM from both species. Replacement of the putative P1′–P2′ residues in mouse NPM with the corresponding residues of human NPM restored cleavage of mouse NPM by both granzymes. This further demonstrates the importance of prime sites as structural determinants for GrM substrate specificity. GrM from both species efficiently triggered apoptosis in human but not in mouse tumour cells. These results indicate that hGrM and mGrM not only exhibit divergent specificities but also trigger species-specific functions.

scholarly journals Mouse and Human Granzyme B Have Distinct Tetrapeptide Specificities and Abilities to Recruit the Bid Pathway

2006 ◽  
Vol 282 (7) ◽  
pp. 4545-4552 ◽  
Author(s):  
Livia Casciola-Rosen ◽  
Margarita Garcia-Calvo ◽  
Herbert G. Bull ◽  
Joseph W. Becker ◽  
Tonie Hines ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Granzyme B ◽  
Target Cells ◽  
Cytotoxic Lymphocyte ◽  
Induce Cell Death ◽  
Important Mediator ◽  
Protease Substrate ◽  
Species Specific ◽  
And Function ◽  
Human And Mouse

Granzyme B is an important mediator of cytotoxic lymphocyte granule-induced death of target cells, accomplishing this through cleavage of Bid and cleavage and activation of caspases as well as direct cleavage of downstream substrates. Significant controversy exists regarding the primary pathways used by granzyme B to induce cell death, perhaps arising from the use of different protease/substrate combinations in different studies. The primary sequence of human, rat, and mouse granzymes B is well conserved, and the substrate specificity and crystal structure of the human and rat proteases are extremely similar. Although little is known about the substrate specificity of mouse granzyme B, recent studies suggest that it may differ significantly from the human protease. In these studies we show that the specificities of human and mouse granzymes B differ significantly. Human and mouse granzyme B cleave species-specific procaspase-3 more efficiently than the unmatched substrates. The distinct specificities of human and mouse granzyme B highlight a previously unappreciated requirement for Asp192 in the acquisition of catalytic activity upon cleavage of procaspase-3 at Asp175. Although human granzyme B efficiently cleaves human or mouse Bid, these substrates are highly resistant to cleavage by the mouse protease, strongly indicating that the Bid pathway is not a major primary mediator of the effects of mouse granzyme B. These studies provide important insights into the substrate specificity and function of the granzyme B pathway in different species and highlight that caution is essential when designing and interpreting experiments with different forms of granzyme B.

Synthesis of positional-scanning libraries of fluorogenic peptide substrates to define the extended substrate specificity of plasmin and thrombin

10.1038/72642 ◽  
2000 ◽  
Vol 18 (2) ◽  
pp. 187-193 ◽  
Author(s):  
Bradley J. Backes ◽  
Jennifer L. Harris ◽  
Francesco Leonetti ◽  
Charles S. Craik ◽  
Jonathan A. Ellman
Keyword(s):  
Substrate Specificity ◽  
Peptide Substrates ◽  
Positional Scanning ◽  
Fluorogenic Peptide ◽  
Positional Scanning Libraries

scholarly journals Human cytotoxic lymphocyte tryptase. Its purification from granules and the characterization of inhibitor and substrate specificity.

1988 ◽  
Vol 263 (26) ◽  
pp. 13215-13222 ◽  
Author(s):  
M Poe ◽  
C D Bennett ◽  
W E Biddison ◽  
J T Blake ◽  
G P Norton ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Cytotoxic Lymphocyte

Investigation of Structural Determinants for the Substrate Specificity in the Zinc-Dependent Alcohol Dehydrogenase CPCR2 fromCandida parapsilosis

ChemBioChem ◽  
2015 ◽  
Vol 16 (10) ◽  
pp. 1512-1519 ◽  
Author(s):  
Christoph Loderer ◽  
Gaurao V. Dhoke ◽  
Mehdi D. Davari ◽  
Wolfgang Kroutil ◽  
Ulrich Schwaneberg ◽  
...  
Keyword(s):  
Alcohol Dehydrogenase ◽  
Substrate Specificity ◽  
Structural Determinants

Identification of Novel Ligands for Vasopressin V2 Receptor from Positional Scanning Libraries

2001 ◽  
pp. 921-922
Author(s):  
Colette T. Dooley ◽  
Aric Mabini ◽  
Tyler Sylvester ◽  
Jon R. Appel ◽  
Richard A. Houghten
Keyword(s):  
Vasopressin V2 Receptor ◽  
Positional Scanning ◽  
V2 Receptor ◽  
Positional Scanning Libraries

scholarly journals Are terrestrial isopods able to use stridulation and vibrational communication as forms of intra and interspecific signaling and defense strategies as insects do? A preliminary study in Armadillo officinalis

2019 ◽  
Vol 107 (1) ◽  
Author(s):  
Sofia Cividini ◽  
Spyros Sfenthourakis ◽  
Giuseppe Montesanto
Keyword(s):  
Defense Mechanism ◽  
High Sensitivity ◽  
Specific Substrate ◽  
Test Apparatus ◽  
Defense Strategies ◽  
Terrestrial Isopods ◽  
Adverse Conditions ◽  
Secondary Form ◽  
Preliminary Study ◽  
Species Specific

AbstractThe capability of producing sounds and vibrations is well known in insects and is thought to be a form of intra- and interspecific communication. Sounds and vibrations are used and modulated for several aims such as interacting with conspecifics, getting information from the environment, and defending against predators. This phenomenon is less known but also present in other arthropods, including a few roller-type terrestrial isopods. In this study, we used a Y-shape test apparatus to investigate the behavior of adult individuals of Armadillo officinalis Duméril, 1816 (Crustacea: Isopoda: Oniscidea) when exposed to two particular vibrational stimuli, namely species-specific stridulations and non-specific substrate-borne vibrations. Our results showed that adults of A. officinalis significantly react to the presence of both types of vibrational stimuli, by moving away from the vibrational source as if they experienced these vibrations as a sign of danger or disturbance. A. officinalis can produce stridulations only when it rolls into a ball during the so-called conglobation, a possible defense mechanism against predators. Stridulation might thus be a secondary form of defense used during conglobation to deter a predator following contact with it and might be experienced as an alert by conspecifics nearby. The high sensitivity to non-specific substrate-borne vibrations might provide A. officinalis with the possibility to anticipate dangers and adverse conditions, giving it a better chance of survival.

scholarly journals Activity profiling and crystal structures of inhibitor-bound SARS-CoV-2 papain-like protease: A framework for anti–COVID-19 drug design

2020 ◽  
Vol 6 (42) ◽  
pp. eabd4596 ◽  
Author(s):  
Wioletta Rut ◽  
Zongyang Lv ◽  
Mikolaj Zmudzinski ◽  
Stephanie Patchett ◽  
Digant Nayak ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Crystal Structures ◽  
Drug Repurposing ◽  
Activity Profiling ◽  
Irreversible Inhibitors ◽  
Inhibitory Mechanisms ◽  
Promising Target ◽  
Therapeutic Value ◽  
Positional Scanning ◽  
High Degree

Viral papain-like cysteine protease (PLpro, NSP3) is essential for SARS-CoV-2 replication and represents a promising target for the development of antiviral drugs. Here, we used a combinatorial substrate library and performed comprehensive activity profiling of SARS-CoV-2 PLpro. On the scaffold of the best hits from positional scanning, we designed optimal fluorogenic substrates and irreversible inhibitors with a high degree of selectivity for SARS PLpro. We determined crystal structures of two of these inhibitors in complex with SARS-CoV-2 PLpro that reveals their inhibitory mechanisms and provides a molecular basis for the observed substrate specificity profiles. Last, we demonstrate that SARS-CoV-2 PLpro harbors deISGylating activity similar to SARSCoV-1 PLpro but its ability to hydrolyze K48-linked Ub chains is diminished, which our sequence and structure analysis provides a basis for. Together, this work has revealed the molecular rules governing PLpro substrate specificity and provides a framework for development of inhibitors with potential therapeutic value or drug repurposing.

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