重组 PPi 依赖的 6-磷酸果糖激酶的表征来自甲基营养型沼泽红假单胞菌 OB3b 和甲基杆菌 nodulans ORS 2060。

Characterization of recombinant PPi-dependent 6-phosphofructokinases from Methylosinus trichosporium OB3b and Methylobacterium nodulans ORS 2060.

机构信息

Skryabin Institute of Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences, Pushchino, Moscow Region, 142290, Russia.

出版信息

Biochemistry (Mosc). 2012 Mar;77(3):288-95. doi: 10.1134/S0006297912030078.

DOI:10.1134/S0006297912030078

PMID:22803946

Abstract

The properties of the purified recombinant PPi-dependent 6-phosphofructokinases (PPi-PFKs) from the methanotroph Methylosinus trichosporium OB3b and rhizospheric phytosymbiont Methylobacterium nodulans ORS 2060 were determined. The dependence of activities of PPi-PFK-His(6)-tag from Ms. trichosporium OB3b (6 × 45 kDa) and PPi-PFK from Mb. nodulans ORS 2060 (4 × 43 kDa) on the concentrations of substrates of forward and reverse reactions conformed to Michaelis-Menten kinetics. Besides fructose-6-phosphate, the enzymes also phosphorylated sedoheptulose-7-phosphate. ADP or AMP (1 mM each) inhibited activity of the Ms. trichosporium PPi-PFK but did not affect the activity of the Mb. nodulans enzyme. Preference of PPi-PFKs to fructose-1,6-bisphosphate implied a predominant function of the enzymes in hexose phosphate synthesis in these bacteria. PPi-PFKs from the methylotrophs have low similarity of translated amino acid sequences (17% identity) and belong to different phylogenetic subgroups of type II 6-phosphofructokinases. The relationship of PPi-PFKs with microaerophilic character of Ms. trichosporium OB3b and adaptation of Mb. nodulans ORS 2060 to anaerobic phase of phytosymbiosis are discussed.

摘要

从甲烷营养菌 Methylosinus trichosporium OB3b 和根际植物共生菌 Methylobacterium nodulans ORS 2060 中纯化的依赖于焦磷酸的 6-磷酸果糖激酶（PPi-PFK）的性质已被确定。M. trichosporium OB3b 的 PPi-PFK-His（6）-标签（6×45 kDa）和 Mb. nodulans ORS 2060 的 PPi-PFK（4×43 kDa）的活性对正向和反向反应的底物浓度的依赖性符合米氏动力学。除了果糖-6-磷酸外，这些酶还磷酸化景天庚酮糖-7-磷酸。ADP 或 AMP（各 1 mM）抑制 Ms. trichosporium PPi-PFK 的活性，但不影响 Mb. nodulans 酶的活性。PPi-PFK 对果糖-1,6-双磷酸的偏好表明这些酶在这些细菌中六碳磷酸合成中的主要功能。甲基营养菌的 PPi-PFK 具有较低的翻译氨基酸序列相似性（17%的同一性），并属于 II 型 6-磷酸果糖激酶的不同系统发育亚群。讨论了 PPi-PFK 与 Ms. trichosporium OB3b 的微需氧特性以及 Mb. nodulans ORS 2060 对植物共生厌氧阶段的适应之间的关系。