PMID- 35934298
OWN - NLM
STAT- MEDLINE
DCOM- 20220823
LR  - 20230902
IS  - 1878-1454 (Electronic)
IS  - 1570-9639 (Print)
IS  - 1570-9639 (Linking)
VI  - 1870
IP  - 9
DP  - 2022 Sep 1
TI  - Arg236 in human chymotrypsin B2 (CTRB2) is a key determinant of high enzyme 
      activity, trypsinogen degradation capacity, and protection against pancreatitis.
PG  - 140831
LID - S1570-9639(22)00078-4 [pii]
LID - 10.1016/j.bbapap.2022.140831 [doi]
AB  - Pancreatic chymotrypsins (CTRs) are digestive proteases that in humans include 
      CTRB1, CTRB2, CTRC, and CTRL. The highly similar CTRB1 and CTRB2 are the products 
      of gene duplication. A common inversion at the CTRB1-CTRB2 locus reverses the 
      expression ratio of these isoforms in favor of CTRB2. Carriers of the inversion 
      allele are protected against the inflammatory disorder pancreatitis presumably 
      via their increased capacity for CTRB2-mediated degradation of harmful 
      trypsinogen. To reveal the protective molecular determinants of CTRB2, we 
      compared enzymatic properties of CTRB1, CTRB2, and bovine CTRA (bCTRA). By 
      evolving substrate-like Schistocerca gregaria proteinase inhibitor 2 (SGPI-2) 
      inhibitory loop variants against the chymotrypsins, we found that the substrate 
      binding groove of the three enzymes had overlapping specificities. Based on the 
      selected sequences, we produced eight SGPI-2 variants. Remarkably, CTRB2 and 
      bCTRA bound these inhibitors with significantly higher affinity than CTRB1. 
      Moreover, digestion of peptide substrates, beta casein, and human anionic 
      trypsinogen unequivocally confirmed that CTRB2 is a generally better enzyme than 
      CTRB1 while the potency of bCTRA lies between those of the human isoforms. 
      Unexpectedly, mutation D236R alone converted CTRB1 to a CTRB2-like high activity 
      protease. Modeling indicated that in CTRB1 Met210 partially obstructed the 
      substrate binding groove, which was relieved by the D236R mutation. Taken 
      together, we identify CTRB2 Arg236 as a key positive determinant, while CTRB1 
      Asp236 as a negative determinant for chymotrypsin activity. These findings 
      strongly support the concept that in carriers of the CTRB1-CTRB2 inversion 
      allele, the superior trypsinogen degradation capacity of CTRB2 protects against 
      pancreatitis.
CI  - Copyright (c) 2022 The Authors. Published by Elsevier B.V. All rights reserved.
FAU - Nemeth, Balint Zoltan
AU  - Nemeth BZ
AD  - Department of Biochemistry, ELTE Eotvos Lorand University, Pazmany Peter setany 
      1/C, H-1117 Budapest, Hungary.
FAU - Demcsak, Alexandra
AU  - Demcsak A
AD  - Department of Surgery, University of California Los Angeles, Los Angeles, 
      California 90095, USA.
FAU - Micsonai, Andras
AU  - Micsonai A
AD  - ELTE NAP Neuroimmunology Research Group, Department of Biochemistry, ELTE Eotvos 
      Lorand University, Pazmany Peter setany 1/C, H-1117 Budapest, Hungary.
FAU - Kiss, Bence
AU  - Kiss B
AD  - Department of Biochemistry, ELTE Eotvos Lorand University, Pazmany Peter setany 
      1/C, H-1117 Budapest, Hungary.
FAU - Schlosser, Gitta
AU  - Schlosser G
AD  - Department of Analytical Chemistry, MTA-ELTE Lendulet Ion Mobility Mass 
      Spectrometry Research Group, Institute of Chemistry, ELTE Eotvos Lorand 
      University, Pazmany Peter setany 1/A, H-1117 Budapest, Hungary.
FAU - Geisz, Andrea
AU  - Geisz A
AD  - Department of Molecular and Cell Biology, Boston University, Henry M. Goldman 
      School of Dental Medicine, Boston, MA 02118, USA.
FAU - Hegyi, Eszter
AU  - Hegyi E
AD  - Institute for Translational Medicine, University of Pecs, Medical School, Pecs, 
      Hungary.
FAU - Sahin-Toth, Miklos
AU  - Sahin-Toth M
AD  - Department of Surgery, University of California Los Angeles, Los Angeles, 
      California 90095, USA.
FAU - Pal, Gabor
AU  - Pal G
AD  - Department of Biochemistry, ELTE Eotvos Lorand University, Pazmany Peter setany 
      1/C, H-1117 Budapest, Hungary. Electronic address: gabor.pal@ttk.elte.hu.
LA  - eng
GR  - R01 DK082412/DK/NIDDK NIH HHS/United States
GR  - R01 DK117809/DK/NIDDK NIH HHS/United States
PT  - Journal Article
DEP - 20220805
PL  - Netherlands
TA  - Biochim Biophys Acta Proteins Proteom
JT  - Biochimica et biophysica acta. Proteins and proteomics
JID - 101731734
RN  - 0 (Peptides)
RN  - 9002-08-8 (Trypsinogen)
RN  - EC 3.4.21.1 (Chymotrypsin)
RN  - EC 3.4.21.1 (chymotrypsin B)
SB  - IM
MH  - Animals
MH  - Cattle
MH  - *Chymotrypsin/genetics
MH  - Humans
MH  - Pancreas/metabolism
MH  - *Pancreatitis/genetics
MH  - Peptides/metabolism
MH  - Trypsinogen/genetics
PMC - PMC9426946
MID - NIHMS1831812
OTO - NOTNLM
OT  - Directed protein evolution
OT  - Human chymotrypsin
OT  - Pancreatitis
OT  - Phage display
OT  - Serine protease
OT  - Serine protease inhibitor
COIS- CONFLICT OF INTEREST The authors declare that they have no conflict of interest 
      with the contents of this article.
EDAT- 2022/08/08 06:00
MHDA- 2022/08/24 06:00
PMCR- 2023/09/01
CRDT- 2022/08/07 19:38
PHST- 2022/06/10 00:00 [received]
PHST- 2022/07/29 00:00 [revised]
PHST- 2022/08/02 00:00 [accepted]
PHST- 2022/08/08 06:00 [pubmed]
PHST- 2022/08/24 06:00 [medline]
PHST- 2022/08/07 19:38 [entrez]
PHST- 2023/09/01 00:00 [pmc-release]
AID - S1570-9639(22)00078-4 [pii]
AID - 10.1016/j.bbapap.2022.140831 [doi]
PST - ppublish
SO  - Biochim Biophys Acta Proteins Proteom. 2022 Sep 1;1870(9):140831. doi: 
      10.1016/j.bbapap.2022.140831. Epub 2022 Aug 5.