Abstract
THE METABOLISM OF HYDROXYPROLINE IN THE INTACT RAT. INCORPORATION OF HYDROXYPROLINE INTO PROTEIN AND URINARY METABOLITES
Highlights
2000-fold dilution over the fed hydroxyproline. When this is compared to the g-fold dilution of the specific activity in free liver hydroxyproline, it is evident that the newly formed protein-bound hydroxyproline could not have been derived from the free hydroxyproline and that it must have been produced in the bound state
By determining the specific activity of hydroxyproline in carcass protein, as well as the amount of total hydroxyproline, it was possible to calculate the specific activity of hydroxyproline in protein newly synthesized during the experimental period
That the newly formed, protein-bound hydroxyproline could not have been derived from the free hydroxyproline, but must have been produced in the bound state
Summary
The classical studies of Stetten (1)) with N15-labeled proline and hydroxyproline, led to two important conclusions in regard to incorporation of hydroxyproline into protein: Very little hydroxyproline, when fed, is incorporated into protein, most of the tissue hydroxyproline being derived from proline, and there is no free hydroxyproline pool in the organism, most of the tissue hydroxyproline being formed from proline in the bound state. Liver protein of Rat I was hydrolyzed for 24 hours with 6 N HCl. Total radioactivity of the hydrolysate was determined, and an aliquot (5 per cent of the total) was chromatographed on filter paper sheets as described for the free amino acids. Spot 3 (RF in phenol-water, 0.73; in lutidine-water, 0.66) was identified as pyrrole-2-carboxylic acid by the color reaction (purple) with the dimethylaminobenzaldehyde spray, cochromatography with an authentic sample (S), and the carrier purification and degradation described below. The ether was replaced by water, in which the PCA dissolved on warming and from which it crystallized on cooling (yield, 101.7 mg.) It was recrystallized from ethanol and assayed for percentage of carbon and radioactivity by combustion to carbon dioxide (3). The mercury derivative of pyrrole (9), which remained in the tube, was filtered, washed with water, OF HYDROXYPBOLINE and assayed for percentage of carbon and radioactivity; calculated for (GH4N)zHg*4HgC12, C 6.6 per cent; found, C 5.8 per cent
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