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Characterisation of wild‐type and mutant forms of human monoamine oxidase A and B expressed in a mammalian cell line
Author(s) -
Gottowik Jürgen,
Cesura Andrea M.,
Malherbe Parichehr,
Lang Gabrielle,
Da Prada Mosé
Publication year - 1993
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(93)81512-x
Subject(s) - monoamine oxidase b , biochemistry , monoamine oxidase , enzyme , monoamine oxidase a , d amino acid oxidase , amino acid , chemistry , cysteine , wild type , mutant , isozyme , biology , stereochemistry , microbiology and biotechnology , oxidase test , gene
Monoamine oxidase (MAO)‐A and MAO‐B are FAD‐containing mitochondrial enzymes which catabolize biogenic and xenobiotic amines. The N‐terminal regions of both forms of MAO contain an ADP‐binding consensus sequence found in several dinucleotide‐dependent enzymes, but otherwise show remarkable sequence differences. In order to investigate whether the N‐terminal region of MAOs participates in the different catalytic properties and inhibitor specificities exhibited by MAO‐A and MAO‐B, we constructed chimeric A/B forms and expressed them in a human embryonic kidney cell line (293 cells). The MAO‐A chimeric form containing the N‐terminus (36 amino acids) of MAO‐B and the B chimera having the first 45 amino acid sequence of MAO‐A were both catalytically active. Compared to the respective wild‐type form, they did not show any significant difference in their catalytic properties ( K m , k cat ) towards the substrates tested or in their sensitivity towards inhibitors. This indicates that the N‐terminal region of the two isoenzymes is not involved in the different specificities of MAO‐A and MAO‐B. Substitution of Cys‐397 of MAO‐B, i.e. the residue covalently anchoring FAD, with an Ala or a His residue resulted in the total loss of enzymatic activity, suggesting that the covalent coupling of FAD to MAO occurs specifically at the ‐SH group of cysteine.