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Flavin‐dependent N‐hydroxylating enzymes from Mycobacterium smegmatis and Aspergillus fumigatus
Author(s) -
Sobrado Pablo,
Chocklett Wyatt,
Robinson Reeder
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.513.3
Subject(s) - mycobacterium smegmatis , aspergillus fumigatus , biochemistry , enzyme , flavin group , siderophore , cofactor , flavin adenine dinucleotide , virulence , aspergillus nidulans , hydroxylation , chemistry , biology , mycobacterium tuberculosis , microbiology and biotechnology , gene , tuberculosis , mutant , medicine , pathology
Siderophores are iron‐binding molecules required for virulence by several human pathogens. M. tuberculosis and A. fumigatus siderophores contain hydroxamate groups that are essential for iron binding. Hydroxamate‐containing siderophores are synthesized via the hydroxylation of the amino groups on the side chains of lysine and ornithine by flavin‐dependent N‐hydroxylating enzymes. The N6‐lysine hydroxylase from Mycobacterium smegmatis (MbsG) and the N5‐ornithine hydroxylase from Aspergillus fumigatus (SidA) have been shown to be essential for virulence in these organisms and, thus, represent attractive drug targets. Currently, there is no biochemical or structural information about these enzymes. We present the cloning, expression, and characterization of MbsG and SidA. These enzymes are expressed in the active form containing tightly bound FAD. Kinetic characterization with NADPH, NADH, and various amine containing substrates is also presented and we demonstrate that dinucleotide‐induced conformational changes are required to stabilize the peroxyflavin in this family of enzymes. This research was partially funded by grants from the Oak Ridge Associated Universities and the Virginia Academy of Sciences.