Zinc-binding proteins of newborn rat epidermis

Hematoxylin stain was used to demonstrate localization of Zn2+-binding proteins in the skin of the newborn rat. The binding protein showed different solubilities and the zinc binding was dissociated at pH 4.5. Epidermal proteins were extracted stepwise in Tris-HCl buffer containing NaCl at 4 degrees C and 37 degrees C, 1 M potassium phosphate, and 4 M urea. Zn2+-binding protein was obtained from each extract by means of Zn2+ chelate affinity chromatography. We detected elution of histidine-rich protein from the affinity column with a pH 4.5 buffer by 3H-histidine labeling and immunoreactivity to rabbit IgG directed to histidine-rich protein. Amino acid analysis demonstrated that the compositions of the Zn2+-binding proteins soluble in the different extracts not only resemble each other, but are essentially the same as those of histidine-rich protein of the newborn rat skin as previously reported. However, sodium dodecylsulfate gel electrophoresis showed multiple protein bands ranging from Mr less than 25,000 to Mr greater than 200,000. The findings indicate that histidine-rich protein of different forms and solubility exists, in the epidermis and may be involved in the biological function of the cells through metal-binding.