Inhibition of mouse trypsin isoforms by SPINK1 and effect of human pancreatitis-associated mutations.
Pancreatology
; 23(4): 358-366, 2023 Jun.
Article
em En
| MEDLINE
| ID: mdl-37149461
ABSTRACT
Serine protease inhibitor Kazal type 1 (SPINK1) is a trypsin-selective inhibitor protein secreted by the exocrine pancreas. Loss-of-function SPINK1 mutations predispose to chronic pancreatitis through either reduced expression, secretion, or impaired trypsin inhibition. In this study, we aimed to characterize the inhibitory activity of mouse SPINK1 against cationic (T7) and anionic (T8, T9, T20) mouse trypsin isoforms. Kinetic measurements with a peptide substrate, and digestion experiments with ß-casein indicated that the catalytic activity of all mouse trypsins is comparable. Human SPINK1 and its mouse ortholog inhibited mouse trypsins with comparable efficiency (KD range 0.7-2.2 pM), with the sole exception of T7 trypsin, which was inhibited less effectively by the human inhibitor (KD 21.9 pM). Characterization of four chronic pancreatitis-associated human SPINK1 mutations in the context of the mouse inhibitor revealed that the reactive-loop mutations R42N (human K41N) and I43M (human I42M) impaired SPINK1 binding to trypsin (KD 60 nM and 47.5 pM, respectively), whereas mutations D35S (human N34S) and A56S (human P55S) had no impact on trypsin inhibition. Our results confirmed that high-affinity trypsin inhibition by SPINK1 is conserved in the mouse, and the functional consequences of human pancreatitis-associated SPINK1 mutations can be replicated in the mouse inhibitor.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Inibidor da Tripsina Pancreática de Kazal
/
Pancreatite Crônica
Tipo de estudo:
Risk_factors_studies
Limite:
Animals
/
Humans
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article