Pharmacological Activities and Hydrolysis by Peptidases of [Phospho-Ser6]-Bradykinin (pS6-BK)

Assis, Diego M.; Juliano, Luiz; Paschoalin, Thaysa; Kouyoumdjian, Maria; Calixto, João B.; Santos, Robson A.S.; Pertinhez, Thelma A.; Gauthier, Francis; Moreau, Thierry; Blaber, Michael; Juliano, María A.

doi:10.1016/j.bcp.2015.07.033

Cited by 2 publications

(2 citation statements)

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“…10 ). A good example for future study would be phosphorylated amino acids, which have been found to influence caspase activity 53 , human plasma and tissue kallikreins activities 54 , as well as hydrolysis by peptidases of [Phospho-Ser(6)]-bradykinin 55 .…”

Section: Discussionmentioning

confidence: 99%

Highly sensitive and adaptable fluorescence-quenched pair discloses the substrate specificity profiles in diverse protease families

Poręba¹,

Szalek²,

Rut³

et al. 2017

Sci Rep

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Internally quenched fluorescent (IQF) peptide substrates originating from FRET (Förster ResonanceEnergy Transfer) are powerful tool for examining the activity and specificity of proteases, and a variety of donor/acceptor pairs are extensively used to design individual substrates and combinatorial libraries. We developed a highly sensitive and adaptable donor/acceptor pair that can be used to investigate the substrate specificity of cysteine proteases, serine proteases and metalloproteinases. This novel pair comprises 7-amino-4-carbamoylmethylcoumarin (ACC) as the fluorophore and 2,4-dinitrophenyl-lysine (Lys(DNP)) as the quencher. Using caspase-3, caspase-7, caspase-8, neutrophil elastase, legumain, and two matrix metalloproteinases (MMP2 and MMP9), we demonstrated that substrates containing ACC/ Lys(DNP) exhibit 7 to 10 times higher sensitivity than conventional 7-methoxy-coumarin-4-yl acetic acid (MCA)/Lys(DNP) substrates; thus, substantially lower amounts of substrate and enzyme can be used for each assay. We therefore propose that the ACC/Lys(DNP) pair can be considered a novel and sensitive scaffold for designing substrates for any group of endopeptidases. We further demonstrate that IQF substrates containing unnatural amino acids can be used to investigate protease activities/ specificities for peptides containing post-translationally modified amino acids. Finally, we used IQF substrates to re-investigate the P1-Asp characteristic of caspases, thus demonstrating that some human caspases can also hydrolyze substrates after glutamic acid.The irreversible peptide bond hydrolysis of proteins and polypeptides is the most conserved post-translational modification occurring in biochemical pathways in all living organisms 1,2 . This reaction is catalyzed by proteases, which specifically recognize protein targets to control numerous significant biological processes, including cell survival and cell death and the immune response to various pathogens 3 . The selectivity of proteases for binding and subsequently hydrolyzing a selected group of peptides or proteins is termed substrate specificity 4,5 . The increasing number of chemical tools for substrate specificity profiling allows the development of new, more efficient and more selective small molecule substrates 6,7 , inhibitors 8 , and chemical probes 9 , which are useful for the determination of protease activity and the dissection of their physiological functions.Internally quenched fluorescent (IQF) peptide substrates constitute a convenient tool for examining the specificity of the largest group of proteases -endopeptidases 10 . These substrates contain a paired fluorophore (donor) and quencher (acceptor), which are located on opposite sides of the scissile peptide bond 11,12 . If the fluorophore 1

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Section: Discussionmentioning

confidence: 99%

Highly sensitive and adaptable fluorescence-quenched pair discloses the substrate specificity profiles in diverse protease families

Poręba¹,

Szalek²,

Rut³

et al. 2017

Sci Rep

View full text Add to dashboard Cite

show abstract

“…Regulation of proteolysis susceptibility is one mechanism whereby threonine/serine phosphorylation can regulate protein function 17 18 . Recently, the activity and resistance to hydrolysis of S6-phospho-bradykinin (pBk) could be associated not only with the bulky phosphate group but also with a significant flexibility reduction of the peptide in comparison with Bk, as observed by NMR analysis 19 . Phosphorylation is also a mechanism through which some intracellular peptides escape degradation and may act regulating protein interactions 20 .…”

Section: Discussionmentioning

confidence: 99%

A Naturally Occurring Antibody Fragment Neutralizes Infectivity of Diverse Infectious Agents

Polonelli

Ciociola

Elviri

et al. 2016

Sci Rep

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A phosphorylated peptide, named K40H, derived from the constant region of IgMs was detected in human serum by liquid chromatography coupled to high-resolution mass spectrometry. Synthetic K40H proved to exert a potent in vitro activity against fungal pathogens, and to inhibit HIV-1 replication in vitro and ex vivo. It also showed a therapeutic effect against an experimental infection by Candida albicans in the invertebrate model Galleria mellonella. K40H represents the proof of concept of the innate role that naturally occurring antibody fragments may exert against infectious agents, shedding a new light upon the posthumous role of antibodies and opening a new scenario on the multifaceted functionality of humoral immunity.

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Pharmacological Activities and Hydrolysis by Peptidases of [Phospho-Ser6]-Bradykinin (pS6-BK)

Cited by 2 publications

References 54 publications

Highly sensitive and adaptable fluorescence-quenched pair discloses the substrate specificity profiles in diverse protease families

Highly sensitive and adaptable fluorescence-quenched pair discloses the substrate specificity profiles in diverse protease families

A Naturally Occurring Antibody Fragment Neutralizes Infectivity of Diverse Infectious Agents

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