80206-32-2Relevant academic research and scientific papers
Cloning, expression, purification, cofactor requirements, and steady state kinetics of phosphoketolase-2 from Lactobacillus plantarum
Yevenes, Alejandro,Frey, Perry A.
, p. 121 - 127 (2008/09/20)
The genes xpk1 and xpk2(Δ1-21) encoding phosphoketolase-1 and (Δ1-7)-truncated phosphoketolase-2 have been cloned from Lactobacillus plantarum and expressed in Escherichia coli. Both gene-products display phosphoketolase activity on fructose-6-phosphate in extracts. A N-terminal His-tag construct of xpk2(Δ1-21) was also expressed in E. coli and produced active His-tagged (Δ1-7)-truncated phosphoketolase-2 (hereafter phosphoketolase-2). Phosphoketolase-2 is activated by thiamine pyrophosphate (TPP) and the divalent metal ions Mg2+, Mn2+, or Ca2+. Kinetic analysis and data from the literature indicate the activators are MgTPP, MnTPP, or CaTPP, and these species activate by an ordered equilibrium binding pathway, with Me2+TPP binding first and then fructose-6-phosphate. Phosphoketolase-2 accepts either fructose-6-phosphate or xylulose-5-phosphate as substrates, together with inorganic phosphate, to produce acetyl phosphate and either erythrose-4-phosphate or glyceraldehyde-3-phosphate, respectively. Steady state kinetic analysis of acetyl phosphate formation with either substrate indicates a ping pong kinetic mechanism. Product inhibition patterns with erythrose-4-phosphate indicate that an intermediate in the ping pong mechanism is formed irreversibly. Background mechanistic information indicates that this intermediate is 2-acetyl-TPP. The irreversibility of 2-acetyl-TPP formation might explain the overall irreversibility of the reaction of phosphoketolase-2.
NUCLEAR MAGNETIC RESONANCE STUDIES OF D-ERYTHROSE 4-PHOSPHATE IN AQUEOUS SOLUTION. STRUCTURES OF THE MAJOR CONTRIBUTING MONOMERIC AND DIMERIC FORMS
Duke, Colin C.,MacLeod, John K.,Williams, John F.
, p. 1 - 26 (2007/10/02)
N.m.r. studies show that a concentrated (ca. 1.0 M ) aqueous solution of D-erythrose 4-phosphate is composed of an equilibrium mixture of the monomeric aldehyde and hydrated aldehyde forms, which interconvert rapidly, together with a major contribution from three dimeric forms.In dilute solutions (ca. 0.04 M ), the hydrated monomer is predominant and dimeric forms are not detectable at this concentration.The chemical structures and stereochemistry of the three dimers have been elucidated by (1)H- and (13)C n.m.r. spectroscopy of D-erythrose 4-phosphate and its (4,4-2H2) and (3,4,4'-2H3) derivatives, aided by the use of the model compounds, glycolaldehyde, D-glyceraldehyde, and DL-glyceraldehyde 3-phosphate, which also form dimers in concenrated aqueous solution.In some cases, the tert-butyldimethylsilyl derivatives of the model dimeric compounds were prepared and isolated.The two major dimers of D-erythrose 4-phosphate, I and II are asymmetrically substituted 1,3-dioxane and 1,3-dioxolane stuctures, respectively, and dimer III is the α anomer of dimer I.
