EVIDENCE FOR CHANNELING OF INTERMEDIATES IN THE OXIDATIVE PENTOSE-PHOSPHATE PATHWAY BY SOYBEAN AND PEA NODULE EXTRACTS, YEAST EXTRACTS, ANDPURIFIED YEAST ENZYMES
Pm. Debnam et al., EVIDENCE FOR CHANNELING OF INTERMEDIATES IN THE OXIDATIVE PENTOSE-PHOSPHATE PATHWAY BY SOYBEAN AND PEA NODULE EXTRACTS, YEAST EXTRACTS, ANDPURIFIED YEAST ENZYMES, European journal of biochemistry, 246(2), 1997, pp. 283-290
Evidence is presented that intermediates of the oxidative pentose phos
phate pathway (OPPP) are channeled from one pathway enzyme to the next
. CO2 produced from [1-C-14]glucose in the presence of unlabelled path
way intermediates contained much more radioactivity than predicted by
a model in which pathway-produced intermediates are in equilibrium wit
h identical molecules in the bulk phase. This was the case whether glu
cose 6-phosphate (Glc6P), 6-phosphogluconolactone, or 6-phosphoglucona
te was added. Assumptions involved in calculating the amount of (CO2)-
C-14 predicted for free mixing of C-14-labelled and unlabelled interme
diates are discussed, together with the following results. (a) (CO2)-C
-14 production by pea nodules in the presence of 3 mM 6-phosphoglucona
te was higher than in its absence. (b) Apparent chaneling of intermedi
ates was much higher for purified yeast enzymes than for yeast extract
. (c) 6-Phosphogluconate and 6-phosphogluconolactone were channeled be
tween yeast Glc6P dehydrogenase and 6-phosphogluconate dehydrogenase d
espite the absence of 6-phosphogluconolactonase in the purified yeast
enzyme mixture. (d) When purified yeast hexokinase was physically sepa
rated from Glc6P dehydrogenase and 6-phosphogluconate dehydrogenase by
a dialysis membrane, there was no apparent channeling. (e) Poly(ethyl
ene glycol), high salt and detergents had little effect on apparent ch
anneling of OPPP intermediates, which is consistent with a stable comp
lex of enzymes. On the other hand: density gradient centrifugation exp
eriments suggested a more transient interaction between the enzymes. T
aken together, the results support channeling of OPPP pathway intermed
iates.