Comparison of calcification between Bacterionema matruchotii and Actinomyces naeslundii.

The purpose of this research was to examine the requirements for proteolipid initiation of calcification in culture. Proteolipid from a calcifiable microorganism, Bacterionema matruchotii, was compared with proteolipid isolated from a non-calcifiable microorganism, Actinomyces naeslundii. Although A. naeslundii does not calcify in culture, lyophilized cells and proteolipid-containing extracts do initiate apatite formation. A. naeslundii proteolipid (ANN) differs from B. matruchotii (BMN) in concentration, apoprotein polarity, and phospholipid composition. These differences may alter the ability of ANN to nucleate apatite in the intact cell.

Identification and partial purification of a hydrophobic protein component associated with [3H]spiroperidol-binding activity.

The binding activity of radiolabelled neuroleptic drugs has been used to biochemically and pharmacologically characterize the dopamine receptor in brain. An extract which binds [3H]spiroperidol and exhibits stereoselectivity for (+)- and (-)-butaclamol, has been isolated from the calf striatal microsomal fraction. Specific binding activity in the chloroform-methanol extract of this preparation is enhanced over that of the crude homogenate. The highest specific binding of the chloroform methanol extract is associated with the crude phospholipid component which is enriched in hydrophobic proteins and acidic phospholipids. Subfractionation of the crude phospholipid extract by gel filtration (Sephadex LH-20) yields multiple peaks of [3H]spiroperidol binding activity, however four major zones of specific binding activity were detected. These results demonstrating a close association of phospholipids with a dopamine binding site suggest a functional role for proteolipid in receptor recognition and regulation.

Relationship between proteolipids and calcium-phospholipid-phosphate complexes in Bacterionema matruchotii calcification.

Calcium-phospholipid-phosphate complexes (Ca-PL-P) were isolated from calcified and uncalcified Bacterionema matruchotii and its calcified lipid extracts. Similar complexes were absent from the noncalcifying bacterium Actinomyces naeslundii. The majority of the Ca-PL-P complexes were associated with the proteolipid acidic phospholipid component. Ca-PL-P complexes isolated from B. matruchotii and from calcified proteolipid contained phosphatidylinositol, phosphatidylinositol-4-phosphate, phosphatidylinositol-4,5-diphosphate, and phosphatidylserine. They consisted of approximately 52 mole % Ca, 32 mole % organic P, and 15 mole % Pi. During Ca-PL-P extraction from B. matruchotii or its proteolipid-containing calcified lipid extracts, the proteolipid was dissociated and the apoprotein precipitated as fluff at the aqueous-organic solvent interface, thus explaining the failure to detect protein in Ca-PL-P preparations. When the ability of Ca-PL-P complexes and lipid fractions of B. matruchotii to initiate apatite formation from metastable calcium phosphate solution was compared, the yield of hydroxyapatite decreased as follows: Ca-PL-P greater than proteolipid acidic phospholipids greater than proteolipid greater than crude phospholipid greater than total lipids greater than whole cells.

Characterization of calculus matrix calcification nucleator.

The nucleator of dental calculus matrix calcification, in vitro, was analyzed. Attention focused on proteolipid singularity, amino acid composition and related polarity, and phospholipid components. The data were compared to those of the nucleator of Bacterionema matruchotii calcification.

Characterization of Bacterionema matruchotii calcification nucleator.

The nucleator of Bacterionema matruchotii calcification was characterized. Parameters examined were: proteolipid purity and singularity, amino acid composition and relative polarity, phospholipid composition, apoprotein homogeneity, essentiality of the complex for nucleation, and ordered structure. The data fulfill a requirement for comparisons among apatite-nucleating proteolipids.

Pre-apatitic mineral deposition in Bacterionema matruchotii.

A preliminary examination of calcification of Bacterionema matruchotii was undertaken to provide a base-line for future kinetic and mechanistic studies. Specific ion ratios were correlated with crystallography and ultrastructure. Observations include: three consistent X-ray diffraction patterns; no resolution of cellular EMP by electron microscopy; increasing Ca/Mg, Ca/P during calcification.

Proteolipid and bone matrix calcification in vitro.

Proteolipid was demonstrated to contain the nucleator of bone matrix calcification, in vitro. Crude phospholipid extracted from bone matrix was fractionated by gel filtration. A single, protein-containing fraction induced apatite crystallization in a metastable calcium phosphate solution. The fraction was identified as proteolipid. The result supports the validity of a microbiologic analogue for vertebrate calcification.

Acidic lipids associated with the local mechanism of calcificaiton: a review.

Two current areas of research on hard tissues have focused upon acidic lipids associated with the local mechanism of calcification and the presence of matrix vesicles as the loci for initial mineralization. Data which show that acidic phospholipids are a component of the vesicle membrane provide a common denominator between these two areas. Furthermore, studies on vertebrate, microbial and synthetic lipoprotein calcification indicate that acidic phospholipids play a pivotal role in the local mechanism. Through a sequence of initial Ca2++ binding followed by desolvation and ion concentration, the acidic phospholipid rich membrane provides an environment in which calcification can be facilitated. A proposed mechanism includes in additon to Ca2++ binding, the formation of (CaHPO4)2 dimers and their condensation into a Ca9 (PO4)6 unit bound to the membrane. The bound unit functions as a nucleus for the formation of additonal units to form an amorphous calcium phosphate cluster. Conversion to crystalline apatite would require dehydration of the environment, either by mineral build up exceeding the hydrophobic domain or by breakdown of the membrane.

Nucleation of microbiologic calcification by proteolipid.

The component of crude phospholipid responsible for B. matruchotii calcification was isolated. Crude phospholipid, extracted from the microorganism, was separated into five fractions by column chromatography. A single, protein-containing fraction catalyzed apatite formation in a metastable calcium phosphate solution. The nucleating fraction was identified as a proteolipid.