Inhibition Of Crystallization Kinetics Of Calcium Phosphates And Calcium Oxalates By Phosphorylated Ostepontin

To better understand the biomineralization mechanisms of calcium oxalate in plant, we simulate the formation of pathological minerals(such as kidney stones) in human and the effect of peptides, which could compare the biomineralization between plant and animal in order to ascertain the biomineralization mechanisms of the calcium oxalate crystal in plant. Kidney stone is a common disease in urological surgery, of which the most familiar form is calcareous(calcium containing) stones. About eighty percent of kidney stone consist of calcium oxalate and calcium phosphate. The crystallization of which were inhibited by osteopontin present in urine. However, both of the effect and mechanism of phosphorylated OPN on the growth and dissolution kinetics of crystal surface are poorly defined. The study quantify the nucleation and the growth and dissolution kinetics of steps using atomic force microscopy combined with other bulk crystallization technology and protein analysis methods. The main results are summarized as following: 1. OPN peptide inhibits HAP crystallization in a phosphorylation and/or concentration dependent mannerUnder near physiological conditions(p H 7.40, I = 0.15 mol/L), the role of a 14 amino acid segment of osteopontin in inhibiting hydroxyapatite(HAP) nucleation and growth was kinetically monitored by a newly developed method for Ca–P crystallization. OPN peptide dramatically prolongs the induction time with an increase in the number of phosphorylation sites at their lower concentrations(156 nmol/L). The nonphosphorylated peptide segment inhibits the nucleation by prolonging the induction time at a higher concentration(234 nmol/L). Post-translational phosphorylation modifications of OPN peptides plays a critical role in inhibiting HAP crystallization at a given concentration range. The presence of phosphate groups in OPN peptides not only enhances the stability of the calcium phosphate(Ca–P) nanoparticles, but also inhibits the phase transformation from amorphous calcium phosphate(ACP) to the final crystalline phases. These results clearly showed that OPN inhibits HAP crystallization by delaying nucleation and subsequent growth in a phosphorylation and/or concentration dependent manner. 2. Phosphorylated OPN peptides dual control step kinetics and interfacial energy on DCPD(010) surfaceIn situ observations of the interaction of OPN peptide with DCPD(010) surface using atomic force microscopy(AFM) show that phosphorylated OPN peptides specific absorb to the [1 00]Cc step, thereby prevent step movement. Classic C-V model of step pinning could well explained the inhibition of phosphorylated OPN peptides on step movement under different supersaturation. Furthermore, the phosphate side chains of OPN preferentially binding to the [1 00]Cc steps by electrostatic interaction, may alter mineral-water interfacial energies, thus delaying the formation of active steps during growth. While the phosphorylation-deficient form of this segment fails to inhibit DCPD crystallization. This is agreed with the bulk nucleation results. These in vitro results may reveal the dual control of step kinetics and interfacial energy by phosphorylated OPN peptides. 3. Phosphorylated OPN peptides significantly inhibit COM heterogeneous nucleation and aggregationHere, in situ observations of the dissolution kinetics of the [101]Cc, [1 00]Cc and [101 ]Cc steps on the brushite(010) surfaces by oxalate and OPN, two urinary constituents using in situ AFM in a simulated acidic urinary milieu. The results indicate that the inhibition role of phosphorylated OPN peptides on the step retreat rates through step-specific interactions, which reduce the release of calcium ions and this in turn inhibiting the formation of COM nucleation at the expense of brushite crystals. The interface-coupled dissolution-precipitation mechanism and the regulation of natural proteins may have much broader utility for improving our understanding of the inhibition mechanisms of the stone formation.In the study, we use a combination of surface(in situ AFM) and bulk crystallization kinetics methods to investigate the interaction of organic molecule with pathological crystals, which indicates the kinetic processes of calcium phosphate-calcium oxalate pathological mineralization and offers effective clues for understanding the biomineralization mechanisms of calcium oxalate in banana.