期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 277, 期 44, 页码 41954-41959出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M203492200
关键词
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Calbindin D-28k (calbindin) is a member of the calmodulin superfamily of Ca2+-binding proteins. An intracellular target of calbindin was discovered using bacteriophage display. Human recombinant calbindin was immobilized on magnetic beads and used in affinity purification of phage-displayed peptides from a random 12-mer peptide library. One sequence, SYSSLAKYPSHS, was strongly selected both in the presence of Mg2+ and in the presence of Ca2+. Homology search against the protein sequence data base identified a closely similar sequence, ISSIKEKYPSHS, at residues 55-66 in myo-inositol-1(or 4)-monophosphatase (IMPase, EC 3.1.3.25), which constitute a strongly conserved and exposed region in the three-dimensional structure. IMPase is a key enzyme in the regulation of the activity of the phosphatidylinositol-signaling pathway. It catalyzes the hydrolysis of myo-inositol-1(or 4)-monophosphate to form free myo-inositol, maintaining a supply that represents the precursor for inositol phospholipid second messenger signaling systems. Fluorescence spectroscopy showed that isolated calbindin and IMPase interact with an apparent equilibrium dissociation constant, K-D, of 0.9 mum. Both apo and Ca2+-bound calbindin was found to activate IMPase up to 250-fold, depending on the pH and substrate concentration. The activation is most pronounced at conditions that otherwise lead to a very low activity of IMPase, i.e. at reduced pH and at low substrate concentration.
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