Substrate specificity of human kallikrein 6: Salt and glycosaminoglycan activation effects

Pedro Francisco Angelo, Aurelio Resende Lima, Fabiana M. Alves, Sachiko I. Blaber, Isobel A Scarisbrick, Michael Blaber, Luiz Juliano, Maria Aparecida Juliano

Research output: Contribution to journalArticle

85 Citations (Scopus)

Abstract

Human kallikrein 6 (hK6) is abundantly expressed in the central nervous system and is implicated in demyelinating disease. This study provided biochemical data about the substrate specificity and activation of hK6 by glycosaminoglycans and by kosmotropic salts, which followed the Hofmeister series. The screening of fluorescence resonance energy transfer (FRET) peptide families derived from Abz-KLRSSKQ-EDDnp resulted in the finding that Abz-AFRFSQ-EDDnp (where Abz is ortho-aminobenzoic acid and EDDnp is N-[2,4-dinitrophenyl]ethylenediamine)) is the best synthetic substrate described so far for hK6 (kcat/Km = 38,667 s-1mM -1). It is noteworthy that the AFRFS sequence was found as a motif in the amino-terminal domain of seven human ionotropic glutamate receptor subunits. We also examined the hK6 hydrolytic activity on FRET peptides derived from human myelin basic protein, precursor of the Aβ amyloid peptide, reactive center loop of α1-antichymotrypsin, plasminogen, and maturation and inactivation cleavage sites of hK6, which were described earlier as natural substrates for hK6. The best substrates were derived from myelin basic protein. The hK6 maturation cleavage site was poorly hydrolyzed, and no evidence was found to support a two-step self-activation process reported previously. Finally, we assayed FRET peptides derived from sequences that span the cleavage sites for activation of protease-activated receptors (PAR) 1-4, and only the substrate with the PAR 2 sequence was hydrolyzed. These results further supported the hypothesis that hK6 expressed in the central nervous system is involved in normal myelin turnover/demyelination processes, but it is unlikely to self-activate. This report also suggested the possible modulation of ionotropic glutamate receptors and activation of PAR 2 by hK6.

Original languageEnglish (US)
Pages (from-to)3116-3126
Number of pages11
JournalJournal of Biological Chemistry
Volume281
Issue number6
DOIs
StatePublished - Feb 10 2006

Fingerprint

Kallikreins
Substrate Specificity
Glycosaminoglycans
Salts
Chemical activation
Substrates
Fluorescence Resonance Energy Transfer
PAR-2 Receptor
Ionotropic Glutamate Receptors
Peptides
Neurology
Demyelinating Diseases
ortho-Aminobenzoates
PAR-1 Receptor
Central Nervous System
Protein Precursors
Myelin Basic Protein
Plasminogen
Amyloid
Myelin Sheath

ASJC Scopus subject areas

  • Biochemistry

Cite this

Substrate specificity of human kallikrein 6 : Salt and glycosaminoglycan activation effects. / Angelo, Pedro Francisco; Lima, Aurelio Resende; Alves, Fabiana M.; Blaber, Sachiko I.; Scarisbrick, Isobel A; Blaber, Michael; Juliano, Luiz; Juliano, Maria Aparecida.

In: Journal of Biological Chemistry, Vol. 281, No. 6, 10.02.2006, p. 3116-3126.

Research output: Contribution to journalArticle

Angelo, PF, Lima, AR, Alves, FM, Blaber, SI, Scarisbrick, IA, Blaber, M, Juliano, L & Juliano, MA 2006, 'Substrate specificity of human kallikrein 6: Salt and glycosaminoglycan activation effects', Journal of Biological Chemistry, vol. 281, no. 6, pp. 3116-3126. https://doi.org/10.1074/jbc.M510096200
Angelo, Pedro Francisco ; Lima, Aurelio Resende ; Alves, Fabiana M. ; Blaber, Sachiko I. ; Scarisbrick, Isobel A ; Blaber, Michael ; Juliano, Luiz ; Juliano, Maria Aparecida. / Substrate specificity of human kallikrein 6 : Salt and glycosaminoglycan activation effects. In: Journal of Biological Chemistry. 2006 ; Vol. 281, No. 6. pp. 3116-3126.
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