Department of Molecular Biosciences, 1200 Sunnyside Ave, University of Kansas, Lawrence, KS, 66045, USA.
Department of Chemistry and Biochemistry, 3210 N Cramer St, University of Wisconsin-Milwaukee, Milwaukee, WI, 53211, USA.
Arch Biochem Biophys. 2019 Mar 30;664:40-50. doi: 10.1016/j.abb.2019.01.028. Epub 2019 Jan 26.
The hydroxyornithine transformylase from Pseudomonas aeruginosa is known by the gene name pvdF, and has been hypothesized to use N-formyltetrahydrofolate (N-fTHF) as a co-substrate formyl donor to convert N-hydroxyornithine (OHOrn) to N-formyl- N-hydroxyornithine (fOHOrn). PvdF is in the biosynthetic pathway for pyoverdin biosynthesis, a siderophore generated under iron-limiting conditions that has been linked to virulence, quorum sensing and biofilm formation. The structure of PvdF was determined by X-ray crystallography to 2.3 Å, revealing a formyltransferase fold consistent with N-formyltetrahydrofolate dependent enzymes, such as the glycinamide ribonucleotide transformylases, N-sugar transformylases and methionyl-tRNA transformylases. Whereas the core structure, including the catalytic triad, is conserved, PvdF has three insertions of 18 or more amino acids, which we hypothesize are key to binding the OHOrn substrate. Steady state kinetics revealed a non-hyperbolic rate curve, promoting the hypothesis that PvdF uses a random-sequential mechanism, and favors folate binding over OHOrn.
铜绿假单胞菌中的羟鸟氨酸转甲基酶的基因名称为 pvdF，据推测它使用 N-甲酰四氢叶酸（N-fTHF）作为辅助底物供体，将 N-羟鸟氨酸（OHOrn）转化为 N-甲酰-N-羟鸟氨酸（fOHOrn）。PvdF 存在于绿脓菌素生物合成途径中，这是一种在缺铁条件下产生的铁载体，与毒力、群体感应和生物膜形成有关。通过 X 射线晶体学确定了 PvdF 的结构，分辨率为 2.3Å，揭示了一种与 N-甲酰四氢叶酸依赖性酶（如甘氨酰胺核苷酸转甲基酶、N-糖转甲基酶和甲硫氨酰-tRNA 转甲基酶）一致的甲酰转移酶结构。虽然核心结构包括催化三联体是保守的，但 PvdF 有三个以上 18 个氨基酸的插入，我们推测这些插入是结合 OHOrn 底物的关键。稳态动力学显示出非双曲线速率曲线，这促进了 PvdF 使用随机顺序机制的假设，并有利于叶酸结合而不是 OHOrn。
