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Structural and Biophysical Insights into SPINK1 Bound to Human Cationic Trypsin

Nagel, F; Palm, GJ; Geist, N; McDonnell, TCR; Susemihl, A; Girbardt, B; Mayerle, J; ... Delcea, M; + view all (2022) Structural and Biophysical Insights into SPINK1 Bound to Human Cationic Trypsin. International Journal of Molecular Sciences , 23 (7) p. 3468. 10.3390/ijms23073468. Green open access

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Abstract

(1) The serine protease inhibitor Kazal type 1 (SPINK1) inhibits trypsin activity in zymogen granules of pancreatic acinar cells. Several mutations in the SPINK1 gene are associated with acute recurrent pancreatitis (ARP) and chronic pancreatitis (CP). The most common variant is SPINK1 p.N34S. Although this mutation was identified two decades ago, the mechanism of action has remained elusive. (2) SPINK1 and human cationic trypsin (TRY1) were expressed in E. coli, and inhibitory activities were determined. Crystals of SPINK1–TRY1 complexes were grown by using the hanging-drop method, and phases were solved by molecular replacement. (3) Both SPINK1 variants show similar inhibitory behavior toward TRY1. The crystal structures are almost identical, with minor differences in the mutated loop. Both complexes show an unexpected rotamer conformation of the His63 residue in TRY1, which is a member of the catalytic triad. (4) The SPINK1 p.N34S mutation does not affect the inhibitory behavior or the overall structure of the protein. Therefore, the pathophysiological mechanism of action of the p.N34S variant cannot be explained mechanistically or structurally at the protein level. The observed histidine conformation is part of a mechanism for SPINK1 that can explain the exceptional proteolytic stability of this inhibitor.

Type: Article
Title: Structural and Biophysical Insights into SPINK1 Bound to Human Cationic Trypsin
Open access status: An open access version is available from UCL Discovery
DOI: 10.3390/ijms23073468
Publisher version: https://doi.org/10.3390/ijms23073468
Language: English
Additional information: This is an open access article distributed under the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Keywords: pancreas; pancreatitis; Kazal inhibitor; serine protease; N34S; protein–protein interaction; standard mechanism; catalytic triad; crystal structure; surface plasmon resonance (SPR); isothermal titration calorimetry (ITC); molecular dynamics simulations (MDS)
UCL classification: UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science
UCL > Provost and Vice Provost Offices > UCL BEAMS > Faculty of Engineering Science > Dept of Biochemical Engineering
UCL > Provost and Vice Provost Offices > UCL BEAMS
UCL
URI: https://discovery.ucl.ac.uk/id/eprint/10146623
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