Phosphorylated osteopontin peptides suppress crystallization by inhibiting the growth of calcium oxalate crystals.
BACKGROUND
Osteopontin isolated from human urine [uropontin (uOPN)] is a potent inhibitor of calcium oxalate (CaOx) monohydrate (COM) crystallization. However, specific structural features responsible for its effects on CaOx crystallization were not previously known. The present studies were designed to define molecular features responsible for interactions of uOPN with COM crystals and the inhibition of crystallization.


METHODS
Peptides and phosphopeptides with sequences corresponding to potential crystal binding domains within the protein sequence of osteopontin were synthesized. Then the effects of these peptides on COM crystal growth and crystal aggregation were investigated and their secondary structures analyzed.


RESULTS
Growth of COM crystals was inhibited by approximately 50% at 1000 nmol/L concentrations by the two unmodified peptides with the closest clustering of aspartic acid residues. Growth was not inhibited by the other two unmodified peptides, with aspartic residues more evenly distributed within their sequences. Phosphorylation markedly increased inhibition of COM crystal growth, so that each of the four phosphorylated peptides inhibited growth by at least 50% at concentrations of < or =200 nmol/L. Phosphorylation of these peptides did not cause changes in secondary structure that would favor interaction with COM crystal surfaces.


CONCLUSIONS
These studies of synthetic peptides identify molecular features within the osteopontin molecule that contribute to the inhibition of one aspect of COM crystallization. The inhibition of crystal growth induced by phosphorylation appears to result from altered local patterns of charge density, since conformational changes favoring interaction with crystals were not caused by phosphorylation.
Hoyer
J R
JR
Department of Pediatrics, University of Pennsylvania and Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, USA. hoyer@email.chop.edu
Asplin
J R
JR
Otvos
L
L
D002129:Calcium Oxalate / Q000037:antagonists & inhibitors / Q000737:chemistry*; D003460:Crystallization; D006801:Humans; D053495:Osteopontin; D010446:Peptide Fragments / Q000502:physiology*; D010766:Phosphorylation; D012795:Sialoglycoproteins / Q000502:physiology*
D010446:Peptide Fragments; C506895:SPP1 protein, human; D012795:Sialoglycoproteins; D053495:Osteopontin; D002129:Calcium Oxalate