Abstract
S-(N-Methylthiocarbamoyl)-l-cysteine (MtcCys) functions as a suicide substrate for l-methionine γ-lyase (l-methionine methanethiol-lyase (deaminating), EC 4.4.1.11) from Pseudomonas putida: it undergoes the α,β-elimination and causes time-dependent inactivation. The partition ratio is about 800 per inactivation event. However, the enzyme catalyzes a β-replacement reaction producing S-(β-hydroxyethyl)-l-cysteine in the presence of β-mercaptoethanol (50 mM), but is not inactivated. Thus, β-mercaptoethanol probably reacts with an inactivating species derived enzymatically from MtcCys instead of a nucleophilic amino acid residue at the active site that is modified with MtcCys in the absence of β-mercaptoethanol. When the enzyme was incubated with [1,2,3-U-14C]MtcCys, the tetrameric enzyme was labeled with 4 mol of 14C-labeled inactivator. The enzyme inactivated with [14C]MtcCys slowly regains about 50% of the original enzyme activity with a concomitant loss of about 50% of the radioactivity incorporated after dialysis. These results indicate that the enzyme has two different residues modified with MtcCys at the active site, and that one of the two resulting adducts is unstable and the other is stable to solvolysis. The enzyme is inactivated similarly by S-(N-methylcarbamoyl)-l-cysteine. The amino acid also partitions between α,β-elimination and suicidal inactivation. However, S-ethyl-l-cysteine does not inactivate the enzyme. This is probably due to protection of the enzyme by ethanethiol produced from S-ethyl-l-cysteine as observed for β-mercaptoethanol added to the reaction mixture.
Original language | English |
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Pages (from-to) | 54-60 |
Number of pages | 7 |
Journal | Biochimica et Biophysica Acta (BBA)/Protein Structure and Molecular |
Volume | 785 |
Issue number | 1-2 |
DOIs | |
Publication status | Published - 1984 Feb 28 |
Externally published | Yes |
Keywords
- (P. putida)
- MtcCys
- S-(N-methylthiocarbamoyl)-l-cysteine
- Suicide substrate
- l-methionine γ-lyase
ASJC Scopus subject areas
- Biophysics
- Structural Biology
- Biochemistry
- Molecular Biology