P. W. Wilson

1,25-dihydroxyvitamin D stimulation of specific membrane proteins in chick intestine

Vitamin D-deficient chicks were injected intracardially with physiological doses of 1,25-dihydroxycholecalciferol (1,25-(OH)2D3) and the formation of intestinal brush-border proteins was followed in vitro. Within 4 h of receiving the hormone the incorporation of radioactive leucine into at least two proteins in the brush-borders was increased. The apparent molecular weights of these proteins were 45 000 and 84 000. The change in the synthesis of these proteins was followed with time and compared with the concomitant changes in intestinal calcium transport. The relationship of these changes is such that there is a strong possibility that the proteins are involved in calcium absorption.

Some characteristics of new tissue-binding proteins for metabolites of vitamin D other than 1,25-dihydroxyvitamin D

Protein(s) have been found in a wide range of tissues which have a high affinity for 25-hydroxycholecalciferol. Of the tissues examined only erythrocytes do not have this protein. The properties of the protein have been examined and it has been found that the association constatns range from 2 - 10(9) to 5 - 10(9) M-1 and the sedimentation constants between 5.0 and 6.0 S. It was not possible to distinguish the proteins from the different tissues by their S values, mobility on gel electrophoresis or behaviour on ion-exchange chromatography. These techniques were all used, however, to show that the tissue 25-hydroxycholecalciferol binding protein is distinct from the main plasma binding protein for this steroid and from the intestinal 1,25-dihydroxycholecalciferol-binding protein. A protein has been in the plasma of rachitic animals but not of normals, which is apparently indistinguishable from this new tissue 25-hydroxycholecalciferol-binding protein. The steroid specificity of this new binding protein has been shown to be dependent upon a C-25 hydroxyl group, and an intact conjugated double bond system. Possible functions for this protein have been briefly discussed.

Calcium binding activity by chick intestinal brush-border membrane vesicles

Uptake of Ca2+ by vesicle preparation of chick intestinal brush-border membranes was rapid and extensive. With tracer quantities of Ca2+ uptake was complete in 10 min whereas with 2.0 mM Ca2+ maximum uptake by the vesicles occurred after one hour incubation. The maximum concentration of Ca2+ found in the vesicles was four times greater than the external Ca2+ concentration showing that the majority of the Ca2+ was membrane bound. The Ca2+ taken up by the vesicles was probably bound to the vesicles interior since it was not replaced by exposure of loaded vesicles to La3+ (5 mM). The uptake of Ca2+ by the vesicles at different Ca2+ concentrations was analyzed and a high affinity Ca2+ binding site was found with an association constant for Ca2+ of 5×10−5 M. More of these sites were found in the duodenum than the ileum and vitamin D increases the number of these sites.Uptake of Ca2+ by vesicle preparation of chick intestinal brush-border membranes was rapid and extensive. With tracer quantities of Ca2+ uptake was complete in 10 min whereas with 2.0 mM Ca2+ maximum uptake by the vesicles occurred after one hour incubation. The maximum concentration of Ca2+ found in the vesicles was four times greater than the external Ca2+ concentration showing that the majority of the Ca2+ was membrane bound.The Ca2+ taken up by the vesicles was probably bound to the vesicles interior since it was not replaced by exposure of loaded vesicles to La3+ (5 mM). The uptake of Ca2+ by the vesicles at different Ca2+ concentrations was analyzed and a high affinity Ca2+ binding site was found with an association constant for Ca2+ of 5×10−5 M. More of these sites were found in the duodenum than the ileum and vitamin D increases the number of these sites.