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Biochemistry (Moscow)

, Volume 77, Issue 3, pp 288–295 | Cite as

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

  • O. N. Rozova
  • V. N. KhmeleninaEmail author
  • Y. A. Trotsenko
Article

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-His6-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.

Key words

pyrophosphate-dependent 6-phosphofructokinase methylotrophic bacteria sedoheptulose-1,7-bisphosphatase glycolysis gluconeogenesis 

Abbreviations

ATP-PFK

ATP-dependent 6-phosphofructokinase

FBP

fructose-1,6-bisphosphate

F6P

fructose-6-phosphate

IPTG

isopropyl β-D-thiogalactopyranoside

LB

Luria-Bertani medium

PFK

6-phosphofructokinase

PPi-PFK

pyrophosphate-dependent 6-phosphofructokinase

PRK

phosphoribulokinase

RuBisCO

ribulose-1,5-bisphosphatecarboxylase/oxygenase

RuMP

ribulose monophosphate

SBPase

sedoheptulose-1,7-bisphosphatase

S7P

sedoheptulose-7-phosphate

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Copyright information

© Pleiades Publishing, Ltd. 2012

Authors and Affiliations

  • O. N. Rozova
    • 1
  • V. N. Khmelenina
    • 1
    Email author
  • Y. A. Trotsenko
    • 1
  1. 1.Skryabin Institute of Biochemistry and Physiology of MicroorganismsRussian Academy of SciencesPushchino, Moscow RegionRussia

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