UNIDIRECTIONAL STEADY-STATE RATES OF CENTRAL METABOLISM ENZYMES MEASURED SIMULTANEOUSLY IN A LIVING PLANT-TISSUE

The unidirectional steady state reaction rates of several enzymes and metabolic fluxes of distinct processes were measured simultaneously in hypoxic maize root tips using two-dimensional phosphorus NMR exchange spectroscopy. A single spectrum monitors ATP synthesis and hydrolysis as well as the activities of four enzymes involved in key pathways of central metabolism: UDP-glucose pyrophosphorylase, phosphoglucomutase , hexose-phosphate isomerase, and enolase, The corresponding unidirect ional reaction rates and net metabolic fluxes were calculated from spe ctral intensities. This method provides a unique picture, at enzyme re solution, of how metabolism reacts in a concerted fashion to changes i n external parameters such as temperature and oxygen concentration, By increasing hypoxia via an increase in temperature, we measured the ex pected increase in glycolysis through enolase activity while total ATP synthesis settled. At the same time, we observed a net flux through p hosphoglucomutase and UDP-glucose pyrophosphorylase toward carbohydrat e synthesis. This result is discussed in relation to the current hypot hesis on the turnover of cell walls and sucrose. This reaction also pr oduces a net flux of pyrophosphate, which is needed by pyrophosphate:f ructose-6-phosphate 1-phosphotransferase to work as a glycolytic enzym e.