PMID- 34396457
OWN - NLM
STAT- MEDLINE
DCOM- 20220128
LR  - 20220218
IS  - 1432-0738 (Electronic)
IS  - 0340-5761 (Print)
IS  - 0340-5761 (Linking)
VI  - 95
IP  - 10
DP  - 2021 Oct
TI  - Alkylation of rabbit muscle creatine kinase surface methionine residues inhibits 
      enzyme activity in vitro.
PG  - 3253-3261
LID - 10.1007/s00204-021-03137-6 [doi]
AB  - Creatine kinase (CK) catalyzes the formation of phosphocreatine from adenosine 
      triphosphate (ATP) and creatine. The highly reactive free cysteine residue in the 
      active site of the enzyme (Cys(283)) is considered essential for the enzymatic 
      activity. In previous studies we demonstrated that Cys(283) is targeted by the 
      alkylating chemical warfare agent sulfur mustard (SM) yielding a thioether with a 
      hydroxyethylthioethyl (HETE)-moiety. In the present study, the effect of SM on 
      rabbit muscle CK (rmCK) activity was investigated with special focus on the 
      alkylation of Cys(283) and of reactive methionine (Met) residues. For 
      investigation of SM-alkylated amino acids in rmCK, micro liquid 
      chromatography-electrospray ionization high-resolution tandem-mass spectrometry 
      measurements were performed using the Orbitrap technology. The treatment of rmCK 
      with SM resulted in a decrease of enzyme activity. However, this decrease did 
      only weakly correlate to the modification of Cys(283) but was conclusive for the 
      formation of Met(70)-HETE and Met(179)-HETE. In contrast, the activity of mutants 
      of rmCK produced by side-directed mutagenesis that contained substitutions of the 
      respective Met residues (Met(70)Ala, Met(179)Leu, and Met(70)Ala/Met(179)Leu) was 
      highly resistant against SM. Our results point to a critical role of the surface 
      exposed Met(70) and Met(179) residues for CK activity.
CI  - (c) 2021. The Author(s).
FAU - Steinritz, Dirk
AU  - Steinritz D
AUID- ORCID: 0000-0002-2073-5683
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany. dirk.steinritz@lrz.uni-muenchen.de.
AD  - Walther-Straub-Institute of Pharmacology and Toxicology, 
      Ludwig-Maximilians-Universitat Munich (LMU), Goethestrasse 33, 80366, Munich, 
      Germany. dirk.steinritz@lrz.uni-muenchen.de.
AD  - Bundeswehr Medical Service Academy, Ingolstadter Strasse 240, 80939, Munich, 
      Germany. dirk.steinritz@lrz.uni-muenchen.de.
FAU - Luling, Robin
AU  - Luling R
AUID- ORCID: 0000-0002-6392-2542
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany.
AD  - Walther-Straub-Institute of Pharmacology and Toxicology, 
      Ludwig-Maximilians-Universitat Munich (LMU), Goethestrasse 33, 80366, Munich, 
      Germany.
FAU - Siegert, Markus
AU  - Siegert M
AUID- ORCID: 0000-0003-1760-7733
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany.
AD  - Proteros Biostructures GmbH, Bunsenstrasse 7a, 82152, Planegg, Germany.
FAU - Muckter, Harald
AU  - Muckter H
AD  - Walther-Straub-Institute of Pharmacology and Toxicology, 
      Ludwig-Maximilians-Universitat Munich (LMU), Goethestrasse 33, 80366, Munich, 
      Germany.
FAU - Popp, Tanja
AU  - Popp T
AUID- ORCID: 0000-0002-9028-3870
AD  - Bundeswehr Institute of Radiobiology, Neuherbergstrasse 11, 80937, Munich, 
      Germany.
FAU - Reinemer, Peter
AU  - Reinemer P
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany.
AD  - AM1 Ventures GmbH, Fasanenstrasse 27a, 81247, Munich, Germany.
FAU - Gudermann, Thomas
AU  - Gudermann T
AD  - Walther-Straub-Institute of Pharmacology and Toxicology, 
      Ludwig-Maximilians-Universitat Munich (LMU), Goethestrasse 33, 80366, Munich, 
      Germany.
FAU - Thiermann, Horst
AU  - Thiermann H
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany.
FAU - John, Harald
AU  - John H
AUID- ORCID: 0000-0002-7126-6709
AD  - Bundeswehr Institute of Pharmacology and Toxicology, Neuherbergstrasse 11, 80937, 
      Munich, Germany.
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20210816
PL  - Germany
TA  - Arch Toxicol
JT  - Archives of toxicology
JID - 0417615
RN  - 0 (Chemical Warfare Agents)
RN  - AE28F7PNPL (Methionine)
RN  - EC 2.7.3.2 (Creatine Kinase, MM Form)
RN  - K848JZ4886 (Cysteine)
RN  - T8KEC9FH9P (Mustard Gas)
SB  - IM
MH  - Alkylation/drug effects
MH  - Animals
MH  - Chemical Warfare Agents/*toxicity
MH  - Chromatography, Liquid
MH  - Creatine Kinase, MM Form/*drug effects/metabolism
MH  - Cysteine/metabolism
MH  - Methionine/*metabolism
MH  - Mustard Gas/*toxicity
MH  - Rabbits
MH  - Spectrometry, Mass, Electrospray Ionization
MH  - Tandem Mass Spectrometry
PMC - PMC8448711
OTO - NOTNLM
OT  - Alkylating agents
OT  - Enzyme activity
OT  - Free cysteine residue
OT  - Hydroxyethylthioethyl
OT  - Mass spectrometry
OT  - Methionine
OT  - Sulfur mustard
COIS- The authors declare no conflict of interest.
EDAT- 2021/08/17 06:00
MHDA- 2022/01/29 06:00
PMCR- 2021/08/16
CRDT- 2021/08/16 06:15
PHST- 2021/07/09 00:00 [received]
PHST- 2021/08/11 00:00 [accepted]
PHST- 2021/08/17 06:00 [pubmed]
PHST- 2022/01/29 06:00 [medline]
PHST- 2021/08/16 06:15 [entrez]
PHST- 2021/08/16 00:00 [pmc-release]
AID - 10.1007/s00204-021-03137-6 [pii]
AID - 3137 [pii]
AID - 10.1007/s00204-021-03137-6 [doi]
PST - ppublish
SO  - Arch Toxicol. 2021 Oct;95(10):3253-3261. doi: 10.1007/s00204-021-03137-6. Epub 
      2021 Aug 16.