Binding of calcium to synthetic peptides containing gamma-carboxyglutamic acid.
The Ca2+ binding properties of various gamma-carboxyglutamic acid (gla)-containing synthetic peptides with counterpart sequences in human protein C were investigated employing potentiometry with a Ca(2+)-selective electrode and titration calorimetric techniques. The shortest peptides, FL(gla)(gla)LR, DF(gla)(gla)AK, and the oxidized form of the cyclic hexapeptide CI(gla)(gla)IC, each of which contains one pair of gla residues, have a weak affinity for Ca2+, with some peptides probably involved in intermolecular bridging of the Ca2+. The best example of this is the oxidized form of the peptide, CI(gla)(gla)IC, where one g-atom of Ca2+ interacts with 2 mol of peptide (n = 0.5) with a Kd value of 1.6 mM. A second g-atom of Ca2+ interacts with 2 mol of this same peptide (n = 0.5) and is characterized by a Kd of 8.8 mM. A longer peptide containing this same sequence, viz. L(gla)R(gla)CI(gla)(gla)IC, possesses two binding sites (n = 2.0) for Ca2+ of Kd = 16.1 mM, as well as a tighter site (n = 1), of Kd = 0.4 mM. An increase in stoichiometry of tight binding sites as the peptide is elongated is observed from binding data obtained on a 38-residue peptide that possesses all nine of the gla-residues of protein C in their proper sequence positions. The strongest Ca2+ binding sites (n = 3-4) possess an average Kd of 0.4 mM, followed by another class of sites (n = 5-10, average Kd = 1.5-3.0 mM). The affinity and stoichiometry of these stronger sites mimic those observed for binding of Ca2+ to the gla region of prothrombin fragment 1.(ABSTRACT TRUNCATED AT 250 WORDS)
Colpitts
T L
TL
Department of Chemistry and Biochemistry, University of Notre Dame, Indiana.
Castellino
F J
FJ
D015055:1-Carboxyglutamic Acid / Q000378:metabolism*; D000595:Amino Acid Sequence; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D002151:Calorimetry; D004566:Electrodes; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010455:Peptides / Q000138:chemical synthesis / Q000378:metabolism*; D011199:Potentiometry; D011486:Protein C / Q000031:analogs & derivatives*; D013329:Structure-Activity Relationship; D017186:Titrimetry
D010455:Peptides; D011486:Protein C; D015055:1-Carboxyglutamic Acid; D002118:Calcium