Des-1-25-fructose-1,6-bisphosphatase, a nonallosteric derivative produced by trypsin treatment of the native protein.
Limited tryptic digestion of pig kidney fructose-1,6-bisphosphatase in the presence of magnesium ions results in the formation of an active enzyme derivative which is no longer inhibited by the allosteric effector AMP. The presence of AMP during incubation of fructose-1,6-bisphosphatase with trypsin protects against the loss of AMP inhibition. By contrast, the presence of the nonhydrolyzable substrate analog fructose 2,6-bisphosphate accelerates the rate of formation of that form of fructose-1,6-bisphosphatase which is insensitive to AMP inhibition. Sodium dodecyl sulfate-polyacrylamide electrophoresis of samples taken during trypsin treatment shows that the loss of AMP inhibition parallels the conversion of the native 36,500 molecular weight fructose-1,6-bisphosphatase subunit into a 34,000 molecular weight species. Automated Edman degradation of trypsin-treated fructose-1,6-bisphosphatase following gel filtration shows a single sequence beginning at Gly-26 in the original enzyme, but no changes in the COOH-terminal region of fructose-1,6-bisphosphatase. Thus, the proteolytic product has been characterized as "des-1-25-fructose-1,6-bisphosphatase." A comparison of the kinetic properties of control enzyme and des-1-25-fructose-1,6-bisphosphatase reveals some differences in properties (pH optimum, Ka for Mg2+, K+ activation, inhibition by fructose 2,6-bisphosphate) between the two enzymes, but none is so striking as the complete loss of AMP sensitivity shown by des-1-25-fructose-1,6-bisphosphatase. The loss of AMP inhibition is due to the loss of AMP-binding capacity, but it is not known at this stage whether residues of the AMP site are present in the 25-amino acid NH2-terminal region or the removal of this region leads to a conformational change that abolishes the function of an AMP site located elsewhere in the molecule.
D000249:Adenosine Monophosphate / Q000494:pharmacology; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D006597:Fructose-Bisphosphatase / Q000032:analysis* / Q000037:antagonists & inhibitors; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D008970:Molecular Weight; D010446:Peptide Fragments / Q000032:analysis*; D011487:Protein Conformation; D051381:Rats; D013552:Swine; D014357:Trypsin / Q000494:pharmacology
D000596:Amino Acids; D010446:Peptide Fragments; D000249:Adenosine Monophosphate; D006597:Fructose-Bisphosphatase; D014357:Trypsin; D008274:Magnesium