Structural studies of a phosphatidyl serine-amorphous calcium phosphate complex

Taylor, Marina G.; Simkiss, K.; Simmons, John R.; Wu, Licia N.Y.; Wuthier, Roy E.

doi:10.1007/s000180050143

Cited by 43 publications

(28 citation statements)

References 20 publications

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“…Ca-P distances (ϳ3.2 Å) similar to those in crystalline calcium phosphate minerals (38). The Ca-P distances indicate Ca 2ϩ coordinates with oxygen atoms of the carboxylate and the phosphodiester bridge of PS, as well as with P i of the ACP component of the CPLX.…”

Section: Discussionmentioning

confidence: 77%

In Vitro Modeling of Matrix Vesicle Nucleation

Genge

Wuthier

2007

Journal of Biological Chemistry

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Annexins A5, A2, and A6 (Anx-A5, -A2, and -A6) are quantitatively major proteins of the matrix vesicle nucleational core that is responsible for mineral formation. Anx-A5 significantly activated the induction and propagation of mineral formation when incorporated into synthetic nucleation complexes made of amorphous calcium phosphate (ACP) and Anx-A5 or of phosphatidylserine (PS) plus ACP (PS-CPLX) and Anx-A5. Incorporation of Anx-A5 markedly shortened the induction time, greatly increasing the rate and overall amount of mineral formed when incubated in synthetic cartilage lymph. Constructed by the addition of Ca 2؉ to PS, emulsions prepared in an intracellular phosphate buffer matched in ionic composition to the intracellular fluid of growth plate chondrocytes, these biomimetic PS-CPLX nucleators had little nucleational activity. However, incorporation of Anx-A5 transformed them into potent nucleators, with significantly greater activity than those made from ACP without PS. The ability of Anx-A5 to enhance the nucleation and growth of mineral appears to stem from its ability to form twodimensional crystalline arrays on PS-containing monolayers. However, some stimulatory effect also may result from its ability to exclude Mg 2؉ and HCO 3 ؊ from nucleation sites. Comparing the various annexins for their ability to activate PS-CPLX nucleation yields the following: avian cartilage Anx-A5 > human placental Anx-A5 > avian liver Anx-A5 > avian cartilage Anx-A6 Ͼ Ͼ cartilage Anx-A2. The stimulatory effect of human placental Anx-A5 and avian cartilage Anx-A6 depended on the presence of PS, since in its absence they either had no effect or actually inhibited the nucleation activity of ACP. Anx-A2 did not significantly enhance mineralization. Matrix vesicles (MVs)2 are extracellular, lipid bilayer-enclosed microstructures released by cells that initiate mineral formation in newly forming bone (1-4). MVs have also been found to initiate ectopic calcification in calcific tendonitis, apatite deposition osteoarthritis, cardiac valve calcification, and atherosclerotic lesions (5-7). MVs isolated from growth plate cartilage by gentle proteolytic digestion and trituration of growth plate tissue (8) retain the ability to induce mineral formation when incubated in synthetic cartilage lymph (SCL) (9). They have been shown to be enriched in several phospholipids, especially phosphatidylserine (PS) (10, 11), a lipid with high affinity for Ca 2ϩ (12,13 MVs contain a detergent-stable core that can induce mineral formation (22, 23). The activity of this nucleational core is inhibited by Zn 2ϩ and is destroyed by treatment with pH 6 citrate buffer. Fourier-transform infrared, NMR, and SDS-PAGE characterization (8,(23)(24)(25) has shown that the nucleational core contains three main components: 1) amorphous calcium phosphate (ACP), 2) membrane-associated CPLX, and, of special interest here, 3) the annexins, lipid-dependent Ca 2ϩ -binding proteins that are especially rich in MVs (8,[25][26][27][28].To elucidate the roles of these components...

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Section: Discussionmentioning

confidence: 77%

In Vitro Modeling of Matrix Vesicle Nucleation

Genge

Wuthier

2007

Journal of Biological Chemistry

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“…However, there is a questionable opinion that ACPs might have an apatitic structure but with a crystal size so small, that they are X-ray amorphous. This is supported by X-ray absorption spectroscopic data (EXAFS) on biogenic and synthetic samples (Harries et al 1986(Harries et al , 1987Taylor et al 1998;Peters et al 2000). On the other hand, it was proposed that the basic structural unit of the precipitated ACPs is a 9.5 Å diameter, roughly spherical cluster of ions with the composition of Ca 9 (PO 4 ) 6 (Fig.…”

Section: Acpmentioning

confidence: 84%

Calcium orthophosphates (CaPO 4 ): Occurrence and properties

Dorozhkin¹

2017

Morphologie

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“…Note the ordering of the assembly of ions by the indicated electrostatic bonds, especially between Ca 2ϩ and the oxygen atoms of the inorganic phosphate and carboxylate of PS. A tabulation of the calculated bond distances for many of the relevant ions is presented in Table 1, where they are compared with values determined by direct analysis of the PS-CPLX using RDF-EXAFS (58).…”

Section: Discussionmentioning

confidence: 99%

“…Recall in our experiments that before addition of Ca 2ϩ , PS was present in the 10 mM pH 7.5 P i buffer. The dominant ionic form of P i would be HPO 4 2Ϫ , which being in great excess would readily form an ion pair with the aliphatic amine of PS (64) ⅐⅐⅐PS-NH 2 ⅐⅐⅐HPO 4 2Ϫ ] 0 , the electrically neutral amine, R-NH 2 , which possesses a pair of unshared electrons, would bond to Ca 2ϩ producing the significantly shorter Ca-N distance indicated by the RDF-EXAFS data (58). Such Ca-N bonding in fact is a key feature in powerful chelating agents such as EDTA.…”

Section: Discussionmentioning

confidence: 99%

“…Our in vitro data, which show a progressive decrease in light scattering during this 10-min period, support the concept that deprotonation of the amine would lead to condensation of the PS-CPLX. In fact, in the discussion of the RDF-EXAFS paper, Taylor et al (58) comment on the close Ca-N distance. Previous MD simulations of only Ca 2ϩ in an aqueous solution containing NH 4 OH show stable Ca-N radial distribution functions within 0.5 ns and yielded maxima positions of 2.7 and 4.6 Å for the first and second shell of hydration (65).…”

Section: Discussionmentioning

confidence: 99%

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Analysis and Molecular Modeling of the Formation, Structure, and Activity of the Phosphatidylserine-Calcium-Phosphate Complex Associated with Biomineralization

Genge

Wuthier

2008

Journal of Biological Chemistry

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The nucleational core of matrix vesicles contains a complex (CPLX) of phosphatidylserine (PS), Ca 2؉ , and inorganic phosphate (P i ) that is important to both normal and pathological calcification. Factors required for PS-CPLX formation and nucleational activity were studied using in vitro model systems and molecular dynamic simulations. Ca 2؉ levels required for and rates of PS-CPLX formation were monitored by light scattering at 340 nm, assessing changes in amount and particle size. Fourier transform infrared spectroscopy was used to explore changes in chemical structure and composition. Washing with pH 5 buffer was used to examine the role of amorphous calcium phosphate in CPLX nucleational activity, which was assessed by incubation in synthetic cartilage lymph with varied pH values. Addition of 4 Ca 2؉ /PS was minimally required to form viable complexes. During the critical first 10-min reaction period, rapid reduction in particle size signaled changes in PS-CPLX structure. Fourier transform infrared spectroscopy revealed increasing mineral phosphate that became progressively deprotonated to PO 4 3؊ . This Ca 2؉ -mediated effect was mimicked in part by increasing the Ca 2؉ /PS reaction ratio. Molecular dynamic simulations provided key insight into initial interactions between Ca 2؉ and P i and the carboxyl, amino, and phosphodiester groups of PS. Deduced interatomic distances agreed closely with previous radial distribution function x-ray-absorption fine structure measurements, except for an elongated Ca 2؉ -N distance, suggesting additional changes in atomic structure during the critical 10-min ripening period. These findings clarify the process of PS-CPLX formation, reveal details of its structure, and provide insight into its role as a nucleator of crystalline calcium phosphate mineral formation. One of the essential features of mineral-inducing matrix vesicles (MV)2 is the presence of a nucleation core (1, 2) that contains lipid-calcium-phosphate complexes (CPLX) capable of inducing mineral formation when incubated in a synthetic cartilage lymph (SCL) (3-6). A critical component of CPLX is the acidic phospholipid, phosphatidylserine (PS) (3,7,8), which is known to have a high affinity for Ca 2ϩ (9,10). PS is largely confined to the inner leaflet of the MV membrane (11), and electron micrographs of calcifying MV show the electrondense calcium phosphate precipitate juxtaposed along the inner leaflet of the MV membrane (12).Discovery of the nucleation core stems from the early finding that acidic phospholipids, especially PS, are complexed with Ca 2ϩ at sites of early mineralization (13,14). PS-Ca 2ϩ -P i complexes (PS-CPLX) are present at the early stages of almost all calcifying tissues as follows: growth plate cartilage (14), tumors (15), bone (16), and especially MV (8). Synthetic PS-CPLX formed in the absence of Mg 2ϩ has been found to be a powerful nucleator that rapidly induces hydroxyapatite (HA) formation when incubated in SCL (3, 4); however, when formed from Mg 2ϩ -and HCO 3 Ϫ -containing P i -r...

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Structural studies of a phosphatidyl serine-amorphous calcium phosphate complex

Cited by 43 publications

References 20 publications

In Vitro Modeling of Matrix Vesicle Nucleation

In Vitro Modeling of Matrix Vesicle Nucleation

Calcium orthophosphates (CaPO 4 ): Occurrence and properties

Analysis and Molecular Modeling of the Formation, Structure, and Activity of the Phosphatidylserine-Calcium-Phosphate Complex Associated with Biomineralization

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