Distal side tryptophan, tyrosine and methionine in catalase-peroxidases are covalently linked in solution.
Distal side tryptophan and tyrosine have been shown to be essential in the catalase but not the peroxidase activity of bifunctional catalase-peroxidases (KatGs). Recently published crystal structures suggest that both residues could be part of a novel adduct including in addition a conserved methionine. A mass spectrometric analysis of the tryptic peptides from recombinant wild-type Synechocystis KatG and the variants Trp122Phe, Tyr249Phe and Met275Ile confirms that this novel adduct really exists in solution and thus may be common to all KatGs. Exchange of either Trp122 or Tyr249 prevents cross-linking, whereas exchange of Met275 still allowed bond formation between Trp122 and Tyr249. It is proposed that the covalent bond between Trp and Tyr may form before that between Tyr and Met. The findings are discussed with respect to the mechanism of cross-linking and its role in KatG catalysis.
Jakopitsch
Christa
C
Institute of Chemistry, University of Natural Resources and Applied Life Sciences, Muthgasse 18, A-1190 Vienna, Austria.
Kolarich
Daniel
D
Petutschnig
Gabriele
G
Furtmüller
Paul Georg
PG
Obinger
Christian
C
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D000458:Cyanobacteria / Q000201:enzymology / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D008715:Methionine / Q000737:chemistry; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics; D012996:Solutions; D021241:Spectrometry, Mass, Electrospray Ionization; D014364:Tryptophan / Q000737:chemistry; D014443:Tyrosine / Q000737:chemistry
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D010446:Peptide Fragments; D012996:Solutions; D014443:Tyrosine; D014364:Tryptophan; D008715:Methionine; D010544:Peroxidases; C081610:catalase-peroxidase, bacteria