Biomedical subjects

B Sabsay

Publications and source records attributed to B Sabsay.

8 recordsLinked to original sources

PubMed1992

Dentin phosphophoryn binding to collagen fibrils.

The interaction of rat incisor phosphophoryn with native turkey tendon collagen fibers has been examined by electron microscopy. The binding of phosphophoryn to the tendon fibril surfaces is quite selective. The phosphophoryn is seen as positively or negatively stained globular particles predominantly at the "e" band in the collagen gap region in transmission electron micrographs of the phosphophoryn-reacted fibrils. The selectivity of binding to the fibrils was obtained in the presence of calcium ions, which bind avidly to phosphophoryn. The specific association of phosphophoryn at the "e" band suggests a possible regulation of mineral deposition within the gap regions of the collagen fibrils.

Animals↗

PubMed1991

Domain structure and sequence distribution in dentin phosphophoryn.

Phosphophoryn (PP) is a protein unique to the mineralized matrix of dentin. It also has a unique composition, with aspartic acid and phosphoserine comprising greater than 85% of all amino acid residues. Because of this unique composition and high content of phosphoserine, it has been difficult to apply direct peptide sequencing procedures effectively. However, to understand its function, and to prepare suitable probes for screening cDNA libraries, some sequence distribution information is required. To this end, using bovine (b) and rat incisor (ri) PPs, partial mild acid hydrolysis has been used to cleave at the aspartic acid residues and generate free amino acids and small peptides. The nature of the released amino acids and peptides has been determined. Peptides have also been generated by limited digestion with trypsin. Some of the peptides have been purified by h.p.l.c. techniques and sequenced. About 90% of the bPP and riPP were resistant to trypsin, and the large resistant fragment was sharply depleted of the non-aspartic acid and non-phosphoserine [(P)Ser] residues. All peptides isolated were acidic, but the remaining residues (other than aspartic acid and serine) appeared to be collected in regions flanking the trypsin-resistant core. These data show directly the presence of regions [Asp]n, [(P)Ser]m and [Asp-(P)Ser-Asp]k as prominent sequence features. A domain structure model is proposed.

Acetates↗

PubMed1991

The isolation and partial characterization of a rat incisor dentin matrix polypeptide with in vitro chondrogenic activity.

In vivo implants of demineralized dentin matrix into muscle induce the formation of bone within the muscle. As with bone matrix implants, the bone induction appears to follow a chondrogenic pathway. Outgrowth cells from explants of neonatal rat muscle respond to bone matrix, in vitro, by expressing a heightened synthesis of sulfated proteoglycans and type II collagen, phenotypic of cartilage. The in vitro cell culture system has been used as an assay to monitor the isolation of the factor responsible for expression of this phenotypic transformation. Soluble proteins extracted from rat incisor dentin matrix during demineralization with EDTA, and not precipitable with 1.0 M CaCl2, were active in the in vitro system. The active extract was fractionated by Sephacryl S-100 chromatography in 6 M guanidine HCl, isoelectric focusing in Immobilines, and by reverse phase high performance liquid chromatography. All fractions were assayed for activity at every stage. The final active fraction from the reverse phase chromatography on a Zorbax Poly-F column was purified to homogeneity, and yielded a single spot on two-dimensional gel electrophoresis. The component, RP-4, had pI 5.4-5.5, and an apparent Mr 6,000-10,000, based on globular protein standards. Maximal activity with respect to both sulfate incorporation into proteoglycan and production of type II collagen was in the 1.0-10 ng/ml range. The RP-4 had a unique amino-terminal amino sequence and was rich in Gly, Pro, Glx, and Ala residues. It was different from transforming growth factor-beta and the bone morphogenetic protein family of proteins in these essential features.

Amino Acid Sequence↗

PubMed1990

Concentration-dependent effects of dentin phosphophoryn in the regulation of in vitro hydroxyapatite formation and growth.

The effect of dentin phosphophoryn on hydroxyapatite formation and growth was studied in an in vitro gelatin gel diffusion system. Phosphophoryn, in low concentrations (0.010-1 microgram/ml) promoted de novo hydroxyapatite formation; at a higher concentration (100 micrograms/ml) in the same system, the dentin matrix protein inhibited hydroxyapatite growth. Similar inhibition of hydroxyapatite growth was seen in solution. The intact phosphophoryn was not essential for either inhibition of seeded growth or promotion of mineralization, since the formic acid degraded protein was comparably effective. Transmission electron microscopy of the precipitates formed at 7 days showed no significant differences in crystallite size distribution in the presence and absence of phosphophoryn. However there was a dose-dependent decrease in the number of mineral clusters formed in the presence of increasing amounts of phosphophoryn, suggesting inhibition of secondary nucleation. These data provide support for the postulated 'multifunctional' role of the dentin phosphoprotein in the mineralization process.

Animals↗

PubMed1990

Rat incisor dentine contains a factor which alters the phenotypic expression and stimulates chondrogenesis in fibroblast-like cells in vitro.

A low molecular weight protein fraction isolated under dissociative conditions during the demineralization of rat incisor dentine has the ability to modulate, in culture, the expression of fibroblast-like cells explanted from neonatal rat muscle. The protein fraction enhances the incorporation of 35S-sulphate into a proteoglycan larger in weight than that produced by the uninduced cells; furthermore it induces the production of type II collagen. These changes take place in the absence of cell proliferation as measured by 3H-thymidine incorporation. The altered fibroblast-like cells form nodules and secrete an abundant extracellular matrix which stains for proteoglycan after 7-9 days in culture. These data show that the dentine matrix does contain a factor which can initiate a mitogenesis-independent alteration in the expression of the muscle-explant outgrowth cells. Those changes are consistent with a shift to a chondrogenic mode.

Animals↗

PubMed1989

Isolation of an acidic protein from cholesterol gallstones, which inhibits the precipitation of calcium carbonate in vitro.

In seeking to identify nucleating/antinucleating proteins involved in the pathogenesis of cholesterol gallstones, a major acidic protein was isolated from each of 13 samples of cholesterol gallstones. After the stones were extracted with methyl t-butyl ether to remove cholesterol, and methanol to remove bile salts and other lipids, they were demineralized with EDTA. The extracts were desalted with Sephadex-G25, and the proteins separated by PAGE. A protein was isolated, of molecular weight below 10 kD, which included firmly-bound diazo-positive yellow pigments and contained 24% acidic, but only 7% basic amino acid residues. The presence of N-acetyl glucosamine suggested that this was a glycoprotein. This protein at concentrations as low as 2 micrograms/ml, but neither human serum albumin nor its complex with bilirubin, inhibited calcium carbonate precipitation from a supersaturated solution in vitro. This protein could be precipitated from 0.15 M NaCl solution by the addition of 0.5 M calcium chloride. Considering that cholesterol gallstones contain calcium and pigment at their centers, and that small acidic proteins are important regulators in other biomineralization systems, this protein seems likely to play a role in the pathogenesis of cholesterol gallstones.

Amino Acids↗

PubMed1989

Characterization of the molecular weights of bovine molar, rat incisor, and other phosphophoryns.

The phosphophoryns show anomalous behavior in solution, and are easily degraded during extraction. They appear in varied forms in the teeth of different species and differ in the teeth of the same species in a developmental and age dependent fashion. This set of properties has made the characterization of the phosphophoryns by biochemical means a difficult and controversial subject. Bovine molar phosphophoryn, bPP, has been characterized in detail by a variety of physical methods, and then compared with the PP from other species by DEAE-HPLC, gel electrophoresis, and immunological cross reactivity. The possible existence of a proPP biosynthetic precursor has been investigated by rat incisor organ culture and examination of the 32P and 14C labeled products. These studies all show marked differences in the Mr values for the PP from teeth of different species, even when they are antigenically cross-reactive.

Animals↗

PubMed1986

Matrix proteins of the teeth of the sea urchin Lytechinus variegatus.

The teeth of the sea urchin Lytechinus variegatus grow continuously. The mineral phase, a high magnesium calcite, grows into single crystals within numerous compartments bounded by an organic matrix deposited by the odontoblasts. Electron microscopic examination of glutaraldehyde-fixed Ethylene Diamine Tetra acetic acid (EDTA) demineralized teeth shows the compartment walls to be organized from multiple layers of cell membrane which might contain cytoplasmic protein inclusions. Proteins extracted during demineralization of unfixed teeth were examined by gel electrophoresis, high performance liquid chromatography, and amino acid analysis. The tooth proteins were acidic, they contained phosphoserine, and they were rich in aspartic acid. By contrast, the proteins of similarly extracted mineralized Aristotle's lantern skeletal elements were nonphosphorylated and were rich in glutamic acid. Vertebrate tooth and bone matrix proteins show similar differences. Surprisingly, an antibody to the principle rat incisor phosphoprotein showed a significant cross-reactivity with the urchin tooth protein, by dot-blot and enzyme-linked immunosorbent assay procedures. Thus, the urchin tooth proteins contain epitope regions similar to those which are phenotypic markers of vertebrate odontoblasts. Whether this is an expression of convergent or divergent evolutionary processes, it is likely that the matrix proteins play a similar role in matrix mineralization. The sea urchin tooth may thus be an excellent model for the study of odontoblast-mediated mineral-matrix relationships.

Amino Acids↗