Baculovirus-expressed vitamin D-binding protein-macrophage activating factor (DBP-maf) activates osteoclasts and binding of 25-hydroxyvitamin D-3 does not influence this activity

Title

Baculovirus-expressed vitamin D-binding protein-macrophage activating factor (DBP-maf) activates osteoclasts and binding of 25-hydroxyvitamin D-3 does not influence this activity

Creator

Swamy N; Ghosh S; Schneider G B; Ray R

Publisher

Journal of Cellular Biochemistry

Date

2001
2001

Description

Vitamin D-binding protein (DBP) is a multi-functional serum protein that is converted to vitamin D-binding protein-macrophage activating factor (DBP-maf) by post-translational modification. DBP-maf is a new cytokine that mediates bone resorption by activating osteoclasts, which are responsible for resorption of bone. Defective osteoclast activation leads to disorders like osteopetrosis, characterized by excessive accumulation of bone mass. Previous studies demonstrated that two nonallelic mutations in the rat with osteopetrosis have independent defects in the cascade involved in the conversion of DBP to DBP-maf. The skeletal defects associated with osteopetrosis are corrected in these mutants with in vivo DBP-maf treatment. This study evaluates the effects of various forms of DBP-maf (native, recombinant, and 25-hydroxyvitamin D-3 bound) on osteoclast function in vitro in order to determine some of the structural requirements of this protein that relate to bone resorbing activities. Osteoclast activity was determined by evaluating pit formation using osteoclasts, isolated from the long bones of newborn rats, incubated on calcium phosphate coated, thin film, Ostologic MultiTest Slides. Incubation of osteoclasts with ex vivo generated native DBP-maf resulted in a dose dependent, statistically significant, activation of the osteoclasts. The activation was similar whether or not the vitamin D binding site of the DBP-maf was occupied. The level of activity in response to DBP-maf was greater than that elicited by optimal doses of other known stimulators (PTH and 1,25(OH)(2)D-3) of osteoclast function. Furthermore, another potent macrophage activating factor, interferon-gamma, had no effect on osteoclast activity. The activated form of a full length recombinant DBP, expressed in E. coli showed no activity in the in vitro assay. Contrary to this finding, baculovirus-expressed recombinant DBP-maf demonstrated significant osteoclast activating activity. The normal conversion of DBP to DBP-maf requires the selective removal of galactose and sialic acid from the third domain of the protein. Hence, the differential effects of the two recombinant forms of DBP-maf is most likely related to glycosylation; E. coli expressed recombinant DBP is non-glycosylated, whereas the baculovirus expressed form is glycosylated. These data support the essential role of glycosylation for the osteoclast activating property of DBP-mai. J. Cell. Biochem. 81:535-546, 2001. (C) 2001 Wiley-Liss, Inc.

Subject

activating factor (DBP-maf); albumin; alpha-fetoprotein; Biochemistry & Molecular Biology; bone resorption; bone resorption; Cell Biology; cells; family; gc-globulin; group-specific component; identification; in-vitro; member; osteoclast activation; recombinant DBP-maf; vitamin D-binding protein (DBP); vitamin D-binding protein macrophage

Format

Journal Article

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Rights

Article information provided for research and reference use only. All rights are retained by the journal listed under publisher and/or the creator(s).

Pages

535-546

Issue

3

Volume

81

Citation

Swamy N; Ghosh S; Schneider G B; Ray R, “Baculovirus-expressed vitamin D-binding protein-macrophage activating factor (DBP-maf) activates osteoclasts and binding of 25-hydroxyvitamin D-3 does not influence this activity,” NEOMED Bibliography Database, accessed September 17, 2021, https://neomed.omeka.net/items/show/7263.

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