Characterization of fructose 6 phosphate phosphoketolases purified from Bifidobacterium species.
Fructose 6 phosphate phosphoketolases (F6PPKs) were purified from Bifidobacterium longum BB536, B. dentium ATCC 27534, B. globosum ATCC 25864, and Bifidobacterium animalis ATCC 25527. Concerning ions (Cu++, Zn++, Ca++, Mg++, Fe++, Co++, Mn++) and common enzyme inhibitors (fructose, ammonium sulfate, iodoacetate, and parachloromercuribenzoic acid), no difference appeared between the enzymes. Cu++, parachloromercuribenzoic acid (pCMB), and mercuric acetate induced high enzymatic inhibition. The study of pCMB demonstrated a noncompetitive inhibition. Additional results showed that the sulfhydryl group was not involved in catalytic reaction. Photooxidation experiments and determination of ionizable group pKas (5.16-7.17) suggested the presence of one or more histidines necessary for the catalytic reaction and explained the inhibition observed with pCMB. In light of the noncompetitive inhibition, this group was not directly involved in substrate binding. Determination of Km demonstrated that the affinities for fructose 6 phosphate in the case of animal and human origin strains were close. In addition, the same enzymatic efficiency (Kcat/Km) was obtained for each strain. The F6PPK activity was regulated by sodium pyrophosphate, ATP, and especially by ADP.
Grill
J P
JP
Université de Nancy I, Institut Henry Tissier, Laboratoire de Chimie Biologique, Vandoeuvre les Nancy, France.
Crociani
J
J
Ballongue
J
J
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000446:Aldehyde-Lyases / Q000187:drug effects / Q000302:isolation & purification* / Q000378:metabolism; D001644:Bifidobacterium / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D008751:Methylene Blue / Q000494:pharmacology; D008970:Molecular Weight; D010755:Organophosphates / Q000378:metabolism; D013045:Species Specificity
D004791:Enzyme Inhibitors; D010755:Organophosphates; C011632:acetyl phosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000446:Aldehyde-Lyases; C065556:fructose-6-phosphate phosphoketolase; D008751:Methylene Blue