Allosteric sites of the large subunit of the spinach leaf ADPglucose pyrophosphorylase.
Pyridoxal-P, an analog of 3-phosphoglycerate, activates spinach leaf ADPglucose pyrophosphorylase. Reductive phosphopyridoxylation of the enzyme yields enzyme less dependent on the presence of activator for activity. Labeled pyridoxal-P is incorporated into both the 51- and 54-kDa subunits of the enzyme. The sequence of the single tryptic labeled peptide of the small subunit has been reported (Morell, M., Bloom, M., and Preiss, J. (1988) J. Biol. Chem. 263, 633-637). Three distinct labeled peptides are isolated from the tryptic digest of the large subunit. 3-Phosphoglycerate inhibits incorporation of pyridoxal-P by 80% into all three large subunit tryptic peptides, whereas inorganic phosphate, the allosteric inhibitor, inhibits incorporation into only one of those peptides. Comparison of the sequences of the labeled tryptic peptides with those of large and small subunits of the enzyme from other species indicate their positions in the primary structure of the spinach enzyme large subunit. The conservation in the amino acid sequences surrounding the lysyl residue in the small subunit and of the lysyl residue in the large subunit in higher plant and in cyanobacterial ADPglucose pyrophosphorylases, which are protected from phosphopyridoxylation by both allosteric effectors, suggests that these lysyl residues are involved in activator binding.
Ball
K
K
Department of Biochemistry, Michigan State University, East Lansing 48824.
Preiss
J
J
D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D001665:Binding Sites; D051599:Glucose-1-Phosphate Adenylyltransferase; D005988:Glyceric Acids / Q000494:pharmacology; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000737:chemistry / Q000378:metabolism*; D011732:Pyridoxal Phosphate / Q000378:metabolism; D018724:Spinacia oleracea / Q000201:enzymology*
D005988:Glyceric Acids; D011732:Pyridoxal Phosphate; C005156:3-phosphoglycerate; D009713:Nucleotidyltransferases; D051599:Glucose-1-Phosphate Adenylyltransferase