Characterization of PlGoxB, a flavoprotein required for cysteine tryptophylquinone biosynthesis in glycine oxidase from Pseudoalteromonas luteoviolacea

• Published in: Archives of biochemistry and biophysics 2019-10, Vol.674, p.108110-108110, Article 108110
• Main Authors: Mamounis, Kyle J., Ma, Zhongxin, Sanchez-Amat, Antonio, Davidson, Victor L.
• Format: Article
• Language: English
• Subjects: Amino Acid Oxidoreductases - chemistry; Amino Acid Oxidoreductases - isolation & purification; Amino Acid Oxidoreductases - metabolism; Catalytic Domain; Flavin; Flavin-Adenine Dinucleotide - chemistry; Flavin-Adenine Dinucleotide - metabolism; Flavoprotein; Flavoproteins - chemistry; Flavoproteins - isolation & purification; Flavoproteins - metabolism; Molecular Docking Simulation; Protein Binding; Protein Stability; Protein-protein interaction; Pseudoalteromonas - enzymology; Quinoprotein
• ISSN: 0003-9861; 1096-0384
• DOI: 10.1016/j.abb.2019.108110

LodA-like proteins are oxidases with a protein-derived cysteine tryptophylquinone (CTQ) prosthetic group. In Pseudoalteromonas luteoviolacea glycine oxidase (PlGoxA), CTQ biosynthesis requires post-translational modifications catalyzed by a modifying enzyme encoded by PlgoxB. The PlGoxB protein was expressed and shown to possess a flavin cofactor. PlGoxB was unstable in solution as it readily lost the flavin and precipitated. PlGoxB precipitation was significantly reduced by incubation with either excess FAD or an equal concentration of prePlGoxA, the precursor protein that is its substrate. In contrast, the mature CTQ-bearing PlGoxA had no stabilizing effect. A homology model of PlGoxB was generated using the structure of Alkylhalidase CmIS. The FAD-binding site of PlGoxB in the model was nearly identical to that of the template structure. The bound FAD in PlGoxB had significant solvent exposure, consistent with the observed tendency to lose FAD. This also suggested that interaction of prePlGoxA with PlGoxB at the exposed FAD-binding site could prevent the observed loss of FAD and subsequent precipitation of PlGoxB. A docking model of the putative PlGoxB-prePlGoxA complex was consistent with these hypotheses. The experimental results and computational analysis implicate structural features of PlGoxB that contribute to its stability and function. •GoxB, the enzyme required for CTQ biosynthesis in the glycine oxidase (GoxA) is a flavoprotein.•GoxB interacts specifically with the GoxA precursor protein that lacks CTQ and not with mature GoxA.•Interaction of GoxB with the precursor of GoxA stabilizes both of these unstable proteins.•A homology model of GoxB reveals structural features that contribute to stability and function.