Abstract
Barley (Hordeum vulgare, cv. Bomi) leaf ADP-glucose pyrophosphorylase (AGP) was purified to near-homogeneity, using ammonium sulfate fractionation and heat treatment as well as ion exchange, hydrophobic, and dye-ligand chromatography. The enzyme was found to be composed of two subunit types of 51 and 54 kDa, which were recognized by the respective rabbit antibodies against the small and/or large subunit of spinach leaf AGP. Substrate kinetics and product inhibition studies, carried out in the direction of ADP-glucose synthesis, suggested a sequential Iso Ordered Bi Bi kinetic mechanism for the enzyme. In addition, inhibition patterns with CrATP, a dead-end inhibitor of ATP-utilizing enzymes, were consistent with ATP (synthesis reaction) and ADP-glucose (pyrophosphorolysis reaction) binding first to different free forms of the enzyme. The AGP was potently activated by 3-phosphoglycerate (PGA) (K alpha of 5 microM) and inhibited by orthophosphate (P(i)), with the ratio of the two effectors playing a major role in modulating AGP activity. At low [PGA]/[P(i)] ratios, P(i) could reverse the activating effect of PGA. However, at the [PGA]/[Pi] ratios of 1.4, or higher, some activating effect of P(i) in the presence of PGA was observed. PGA decreased the Km values for all substrates of AGP (in both the synthesis and pyrophosphorolysis directions). Properties of the barley leaf AGP are compared to those of AGP from barley seed endosperm, which is insensitive to PGA/P(i) regulation.
Highlights
The enzywmaes by 3-phosphoglycerate (PGA), the first stable product of the found to be composed of two subunit types of 51 and 54 kDa, whichwere recognizedby the respectiverabbit antibodies against the small andla/orrge subunit of spinach leaf AGP
It is believed that the PGA/Pi mode of regulation is the key factor in the biosynthesis of starch in plants [1, 10, 11].On the other hand, a recently described AGP from barley seed of ATP-utilizing enzymes, were consistent with ATP endosperm [12] was found to be relatively insensitive to PGA
PGA decreased the K, values of the enzyme, The mechanism was based on the results of for all substrates of AGP.Properties of the bar- exchange studies [9]
Summary
Substrate Kinetics-Substrate kinetics of AGP werecarried out by determination of the rate of ADP-glucose synthesis (assayA) in the reaction mixtures containing varyingconcentrations of ATP and Glu-1-P, in the presence of PGA. The K,,, values for ATP and Glu-1-P, determined with PGA present in assays, were 0.08 and 0.11 mM, respectively (see Table 111), similar to those reported for AGP fromspinach leaves [1,16, 17], maizeseeds [15], potato tubers [13, 14], and barley endosperm [12] (see Table 111).
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
