Defective binding of SPINK1 variants is an uncommon mechanism for impaired trypsin inhibition in chronic pancreatitis.
equilibrium binding assay
pancreatitis
reactive-site peptide bond
trypsin inhibitor
tyrosine sulfation
Journal
The Journal of biological chemistry
ISSN: 1083-351X
Titre abrégé: J Biol Chem
Pays: United States
ID NLM: 2985121R
Informations de publication
Date de publication:
Historique:
received:
15
10
2020
revised:
11
01
2021
accepted:
22
01
2021
pubmed:
31
1
2021
medline:
25
8
2021
entrez:
30
1
2021
Statut:
ppublish
Résumé
The serine protease inhibitor Kazal type 1 (SPINK1) protects the pancreas from intrapancreatic trypsin activation that can lead to pancreatitis. Loss-of-function genetic variants of SPINK1 increase the risk for chronic pancreatitis, often by diminishing inhibitor expression or secretion. Variants that are secreted normally have been presumed to be pathogenic because of defective trypsin inhibition, but evidence has been lacking. Here, we report quantitative studies on the inhibition of human trypsins by wildtype SPINK1 and seven secreted missense variants. We found that tyrosine sulfation of human trypsins weakens binding of SPINK1 because of altered interactions with Tyr43 in the SPINK1 reactive loop. Using authentic sulfated human trypsins, we provide conclusive evidence that SPINK1 variants N34S, N37S, R65Q, and Q68R have unimpaired inhibitory activity, whereas variant P55S exhibits a small and clinically insignificant binding defect. In contrast, rare variants K41N and I42M that affect the reactive-site peptide bond of SPINK1 decrease inhibitor binding by 20,000- to 30,000-fold and three- to sevenfold, respectively. Taken together, the observations indicate that defective trypsin inhibition by SPINK1 variants is an uncommon mechanism in chronic pancreatitis. The results also strengthen the notion that a decline in inhibitor levels explains pancreatitis risk associated with the large majority of SPINK1 variants.
Identifiants
pubmed: 33515547
pii: S0021-9258(21)00115-0
doi: 10.1016/j.jbc.2021.100343
pmc: PMC7949130
pii:
doi:
Substances chimiques
SPINK1 protein, human
0
Trypsin Inhibitor, Kazal Pancreatic
50936-63-5
Trypsin
EC 3.4.21.4
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
100343Subventions
Organisme : NIDDK NIH HHS
ID : R01 DK058088
Pays : United States
Organisme : NIDDK NIH HHS
ID : R01 DK117809
Pays : United States
Informations de copyright
Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.
Déclaration de conflit d'intérêts
Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article.
Références
J Biol Chem. 1974 Apr 10;249(7):2009-15
pubmed: 4856434
Pancreas. 2006 Jul;33(1):104-6
pubmed: 16804421
J Biol Chem. 2020 Mar 13;295(11):3447-3455
pubmed: 32014997
Eur J Hum Genet. 2007 Sep;15(9):936-42
pubmed: 17568390
Dig Dis Sci. 2017 Jul;62(7):1692-1701
pubmed: 28536777
Biochem J. 2009 Feb 15;418(1):155-61
pubmed: 18986305
Hum Mutat. 2017 Aug;38(8):1014-1024
pubmed: 28556356
J Biol Chem. 1973 Oct 25;248(20):7207-14
pubmed: 4795503
J Pediatr Gastroenterol Nutr. 2015 May;60(5):675-9
pubmed: 25383785
Biol Chem Hoppe Seyler. 1990 May;371 Suppl:29-36
pubmed: 2400589
Hum Mutat. 2007 May;28(5):469-76
pubmed: 17274009
Biochem Biophys Res Commun. 2000 Nov 19;278(2):286-9
pubmed: 11097832
Protein Expr Purif. 1998 Mar;12(2):291-4
pubmed: 9518472
Gastroenterology. 2000 Sep;119(3):615-23
pubmed: 10982753
FEBS J. 2006 Nov;273(22):5044-50
pubmed: 17087724
Pancreas. 2012 Mar;41(2):329-30
pubmed: 22343981
Biochem Biophys Res Commun. 1967 Nov 30;29(4):508-14
pubmed: 16496527
Gut. 2014 May;63(5):860-1
pubmed: 24052272
Dig Dis Sci. 1986 Mar;31(3):314-21
pubmed: 2936587
J Mol Biol. 1991 Aug 5;220(3):711-22
pubmed: 1870127
Methods Mol Biol. 2011;705:175-94
pubmed: 21125386
Gastroenterology. 2005 Aug;129(2):696-705
pubmed: 16083722
J Biol Chem. 2006 Apr 28;281(17):11879-86
pubmed: 16505482
J Biol Chem. 2011 Jun 24;286(25):22535-45
pubmed: 21515688
Hum Mutat. 2004 Feb;23(2):205
pubmed: 14722925
FEBS J. 2006 Jul;273(13):2942-54
pubmed: 16759229
Nat Genet. 2000 Jun;25(2):213-6
pubmed: 10835640
J Gastroenterol. 2002;37(11):928-34
pubmed: 12483248
Am J Gastroenterol. 2019 Jun;114(6):974-983
pubmed: 30789418
J Biochem. 1989 Dec;106(6):1059-63
pubmed: 2628423
Gastroenterology. 1984 Apr;86(4):681-92
pubmed: 6698368
Gut. 2009 Apr;58(4):478-80
pubmed: 19299380
Digestion. 2013;87(4):229-39
pubmed: 23751316
Pancreas. 2007 May;34(4):423-8
pubmed: 17446841
Minerva Pediatr. 2013 Dec;65(6):669-72
pubmed: 24217635
Gut. 2017 Dec;66(12):2195-2196
pubmed: 28320769
Gastroenterology. 1981 Mar;80(3):461-73
pubmed: 6969677
PLoS One. 2014 Jul 10;9(7):e102063
pubmed: 25010489
J Biol Chem. 1953 Oct;204(2):797-805
pubmed: 13117856
Genes (Basel). 2017 Oct 10;8(10):
pubmed: 28994706
J Biol Chem. 2003 Dec 5;278(49):48580-9
pubmed: 14507909
Hum Mutat. 2017 Dec;38(12):1619
pubmed: 29091332
J Mol Biol. 1996 Jun 28;259(5):995-1010
pubmed: 8683601
JOP. 2003 Mar;4(2):83-8
pubmed: 12629264
Gut. 2007 Oct;56(10):1433-8
pubmed: 17525091
Gut. 2002 May;50(5):687-92
pubmed: 11950817
Gut. 2009 Apr;58(4):545-9
pubmed: 18978175
Gut. 2006 Aug;55(8):1214
pubmed: 16849362
Hum Mutat. 2017 Dec;38(12):1660-1665
pubmed: 28945313
Pancreas. 2017 Jul;46(6):e54-e55
pubmed: 28609377
J Biol Chem. 2000 Jul 28;275(30):22750-5
pubmed: 10801865
Eur J Biochem. 2003 May;270(9):2047-58
pubmed: 12709065
J Med Genet. 2002 May;39(5):347-51
pubmed: 12011155
Gut. 2016 May;65(5):884-6
pubmed: 26719302