title: Evidence for channeling of intermediates in the oxidative pentose phosphate pathway by soybean and pea nodule extracts, yeast extracts, and purified yeast enzymes.
abstract: Evidence is presented that intermediates of the oxidative pentose phosphate pathway (OPPP) are channeled from one pathway enzyme to the next. CO2 produced from [1-14C]glucose in the presence of unlabelled pathway intermediates contained much more radioactivity than predicted by a model in which pathway-produced intermediates are in equilibrium with identical molecules in the bulk phase. This was the case whether glucose 6-phosphate (Glc6P), 6-phosphogluconolactone, or 6-phosphogluconate was added. Assumptions involved in calculating the amount of 14CO2 predicted for free mixing of 14C-labelled and unlabelled intermediates are discussed, together with the following results. (a) 14CO2 production by pea nodules in the presence of 3 mM 6-phosphogluconate was higher than in its absence. (b) Apparent channeling of intermediates was much higher for purified yeast enzymes than for yeast extract. (c) 6-Phosphogluconate and 6-phosphogluconolactone were channeled between yeast Glc6P dehydrogenase and 6-phosphogluconate dehydrogenase despite the absence of 6-phosphogluconolactonase in the purified yeast enzyme mixture. (d) When purified yeast hexokinase was physically separated from Glc6P dehydrogenase and 6-phosphogluconate dehydrogenase by a dialysis membrane, there was no apparent channeling. (e) Poly(ethylene glycol), high salt and detergents had little effect on apparent channeling of OPPP intermediates, which is consistent with a stable complex of enzymes. On the other hand, density gradient centrifugation experiments suggested a more transient interaction between the enzymes. Taken together, the results support channeling of OPPP pathway intermediates.
authors.affiliation: Biology Department, Washington University, St Louis, MO 63130, USA.
authors.forename: P M
authors.initials: PM
authors.lastname: Debnam
authors.forename: G
authors.initials: G
authors.lastname: Shearer
authors.forename: L
authors.initials: L
authors.lastname: Blackwood
authors.forename: D H
authors.initials: DH
authors.lastname: Kohl
mesh_terms: D002250:Carbon Radioisotopes; D005954:Glucosephosphate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D010084:Oxidation-Reduction; D018532:Pisum sativum / Q000378:metabolism*; D010427:Pentose Phosphate Pathway; D010734:Phosphogluconate Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D010936:Plant Extracts / Q000378:metabolism; D013025:Glycine max / Q000378:metabolism*; D015003:Yeasts / Q000201:enzymology / Q000378:metabolism*
chemical_list: D002250:Carbon Radioisotopes; D010936:Plant Extracts; D010734:Phosphogluconate Dehydrogenase; D005954:Glucosephosphate Dehydrogenase