抄録
Vitamin D3 and its active metabolites being insoluble in water, the transport would take place in an associated form with some kind of proteins. Specific binding proteins were identified in rat plasma and intestinal mucosa by using gel filtration (Sephadex G-200), polyacrylamide gel disc electrophoresis and sucrose density gra- dient ultracentrifugation.
The binding proteins of D3 and 25-HCC were identified in rat plasma. Five protein peaks (designated as peaks 1, 2, 3, 4 and 5 in the elution order) were observed on the gel filtration (2. 4x65 cm). Peaks 1, 2 and 4 were bound with D3 and 25-HCC. However, peaks 2 and 4 were bound with a considerable amount of D3 and peak 4 with 25-HCC. The properties of these peak fractions were also analyzed with poly- acrylamide gel disc electrophoresis and sucrose density gradient ultracentrifugation. D3 was bound with more than five bands of gel stained with acid blue black 10 B. Foul bands of them contained a small amount of carbohydrate and lipid, and a band of them was a-globulin band associated specifically with 25-HCC. The sedi- mentation constants of D3 binding protein ware calculated as 14.5 S, 10.5 S, 8.1 S, 6.2 S and 4.3 S, and the 25-HCC were as 14.5 S, 10.5 S, 6.8 S and 4.3 S. The sedi- mentation constant of the protein, to which D3 was bound specifically, was calculated to be about 8. 1 S. The protein for 25-HCC was 6.8 S.
After intra-abdominal injection of D3-3H to D deficient rat, in the cytoplasmic fraction of intestinal mucosa, three protein peaks appeared upon analysis of Sephadex G-200 column to be bound with radioactivities. This suggested that two other peaks might bind metabolites of D3. After D3-1, 2-3H, 25-HCC-26, 27-3H and 1, 25-DHCC-26, 27-3H were incubated with intestinal cytoplasm in vitro (for 20 minutes, at 2°C), the radioactivity was observed corresponding with peaks 1, 2 and 3 respectively. Each protein binds specifically D3 and its active metabolites, respectively. Sedimentation constants of peaks 1, 2 and 3 were 13S, 8.0-8.6S and 3.5-4. OS upon sucrose density gradient ultracentrifugation.
On the other hand, a single protein fraction (10 S) was identified in the nuclei and bound both D3 and its metabolites. Active metabolites of D3 will be translocated by these proteins in the intestinal cell.
