Biomaterial Database

Affinity labeling of rat serum vitamin D binding protein. 1996

N Swamy, and R Ray

Department of Medicine, Boston University School of Medicine, Massachusetts, USA.

Vitamin D binding protein (DBP) plays an essential role in the vitamin D hormone endocrine system in sequestering vitamin D3 and its metabolites with high affinity, and transporting them to various target organs and tissues. In the present investigation, 25-hydroxyvitamin D3-3 beta-(1,2-epoxypropyl)ether (25-OH-D3-epoxide) and 25-hydroxyvitamin D3-3 beta-bromoacetate (25-OH-D3-BE), synthetic analogs of 25-hydroxyvitamin D3 (25-OH-D3), were developed as affinity alkylating reagents for the covalent modification of the 25-OH-D3-binding site in rat vitamin D binding protein (rDBP). Competitive radioligand binding assays of 25-OH-D3-BE and 25-OH-D3-epoxide with affinity-purified rDBP demonstrated that these analogs displaced 25-hydroxy[26(27)-3H]vitamin D3 (3H-25-OH-D3), specifically bound to rDBP, in a dose-dependent fashion. Incubation of rDBP samples with radiolabeled versions of these analogs, i.e., 3H-25-OH-D3-epoxide and 3H-25-OH-D3-BE, resulted in the covalent labeling of rDBP. This labeling was largely prevented when incubations were carried out in the presence of an excess of 25-OH-D3, the natural ligand for rDBP. Labeling-specificity by these analogs was further demonstrated by the covalent labeling, inhibited by coincubation with a large excess of 25-OH-D3, of a single protein band, upon incubating rat serum Cohn IV fraction with 3H-25-OH-D3-epoxide and 3H-25-OH-D3-BE. Collectively, these results strongly suggested that 3H-25-OH-D3-epoxide and 3H-25-OH-D3-BE covalently modified the 25-OH-D3-binding site in rDBP. The reagents described in this report could be important in mapping the 25-OH-D3-binding pocket in rDBP.

UI	MeSH Term	Description	Entries
D011869	Radioligand Assay	Quantitative determination of receptor (binding) proteins in body fluids or tissue using radioactively labeled binding reagents (e.g., antibodies, intracellular receptors, plasma binders).	Protein-Binding Radioassay,Radioreceptor Assay,Assay, Radioligand,Assay, Radioreceptor,Assays, Radioligand,Assays, Radioreceptor,Protein Binding Radioassay,Protein-Binding Radioassays,Radioassay, Protein-Binding,Radioassays, Protein-Binding,Radioligand Assays,Radioreceptor Assays
D002112	Calcifediol	The major circulating metabolite of VITAMIN D3. It is produced in the LIVER and is the best indicator of the body's vitamin D stores. It is effective in the treatment of RICKETS and OSTEOMALACIA, both in azotemic and non-azotemic patients. Calcifediol also has mineralizing properties.	25-Hydroxycholecalciferol,25-Hydroxyvitamin D 3,25-Hydroxycholecalciferol Monohydrate,25-Hydroxyvitamin D3,Calcidiol,Calcifediol Anhydrous,Calcifediol, (3 alpha,5Z,7E)-Isomer,Calcifediol, (3 beta,5E,7E)-Isomer,Calderol,Dedrogyl,Hidroferol,25 Hydroxycholecalciferol,25 Hydroxycholecalciferol Monohydrate,25 Hydroxyvitamin D 3,25 Hydroxyvitamin D3,Anhydrous, Calcifediol,Monohydrate, 25-Hydroxycholecalciferol
D002762	Cholecalciferol	Derivative of 7-dehydroxycholesterol formed by ULTRAVIOLET RAYS breaking of the C9-C10 bond. It differs from ERGOCALCIFEROL in having a single bond between C22 and C23 and lacking a methyl group at C24.	Vitamin D 3,(3 beta,5Z,7E)-9,10-Secocholesta-5,7,10(19)-trien-3-ol,Calciol,Cholecalciferols,Vitamin D3
D000345	Affinity Labels	Analogs of those substrates or compounds which bind naturally at the active sites of proteins, enzymes, antibodies, steroids, or physiological receptors. These analogs form a stable covalent bond at the binding site, thereby acting as inhibitors of the proteins or steroids.	Affinity Labeling Reagents,Labeling Reagents, Affinity,Labels, Affinity,Reagents, Affinity Labeling
D000818	Animals	Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA.	Animal,Metazoa,Animalia
D001665	Binding Sites	The parts of a macromolecule that directly participate in its specific combination with another molecule.	Combining Site,Binding Site,Combining Sites,Site, Binding,Site, Combining,Sites, Binding,Sites, Combining
D001667	Binding, Competitive	The interaction of two or more substrates or ligands with the same binding site. The displacement of one by the other is used in quantitative and selective affinity measurements.	Competitive Binding
D014809	Vitamin D-Binding Protein	An alpha-globulin found in the plasma of man and other vertebrates. It is apparently synthesized in the liver and carries vitamin D and its metabolites through the circulation and mediates the response of tissue. It is also known as group-specific component (Gc). Gc subtypes are used to determine specific phenotypes and gene frequencies. These data are employed in the classification of population groups, paternity investigations, and in forensic medicine.	Group-Specific Component Globulin,Vitamin D-Binding Globulin,25-HCC-Binding Protein,25-Hydroxycholecalciferol-Binding Protein,25-Hydroxyvitamin D-Binding Protein,Calciferol-Binding Protein,Gc Globulin,Transcalciferin,25 HCC Binding Protein,25 Hydroxycholecalciferol Binding Protein,25 Hydroxyvitamin D Binding Protein,Calciferol Binding Protein,Component Globulin, Group-Specific,Globulin, Gc,Globulin, Group-Specific Component,Globulin, Vitamin D-Binding,Group Specific Component Globulin,Vitamin D Binding Globulin,Vitamin D Binding Protein
D051381	Rats	The common name for the genus Rattus.	Rattus,Rats, Laboratory,Rats, Norway,Rattus norvegicus,Laboratory Rat,Laboratory Rats,Norway Rat,Norway Rats,Rat,Rat, Laboratory,Rat, Norway,norvegicus, Rattus
D066298	In Vitro Techniques	Methods to study reactions or processes taking place in an artificial environment outside the living organism.	In Vitro Test,In Vitro Testing,In Vitro Tests,In Vitro as Topic,In Vitro,In Vitro Technique,In Vitro Testings,Technique, In Vitro,Techniques, In Vitro,Test, In Vitro,Testing, In Vitro,Testings, In Vitro,Tests, In Vitro,Vitro Testing, In