Fig. 2: In vitro biochemical characterization of PyrN-catalyzed N-N formation.
From: Molecular basis of enzymatic nitrogen-nitrogen formation by a family of zinc-binding cupin enzymes
a Domain organization of the di-domain enzyme PyrN. Putative key residues involved in metal ion and ATP binding were also shown, together with the two truncated PyrN constructs used in the following assays. b PyrN-catalyzed tRNA-independent formation of 1. Extracted ion chromatograms (EIC = m/z 514, [M + H]+ ion for Fmoc-1) from the LC-MS analysis of PyrN reaction mixtures were displayed. The different combinations of components contained in the reaction mixtures were indicated next to the corresponding LC-MS traces. c In vitro biochemical assays reveal the essential components to the production of 1. Note: the full reaction (FR) mixtures contain PyrN, ATP, l-Glu, l-N6-OH-Lys (2), and MgCl2 in 40 mM Tris-HCl buffer (pH 8.3). d In vitro biochemical assays of PyrN variants. e In vitro biochemical assays of truncated PyrN constructs that contain only the MetRS-like domain or the cupin domain, as shown in (a).
