Studies on Biotransformation of Lysozyme. II. Tissue Distribution of ¹³¹I-Labeled Lysozyme and Degradation in Kidney after Intravenous Injection in Rats

Abstract

The tissue distribution of TCA-precipitable ¹³¹I following intravenous injection of 0.4 mg lysozyme labeled with ¹³¹I per kg in rats was investigated. The renal concentration of TCA-precipitable ¹³¹I was the highest in all the tissues tested, and the level in the kidney reached the maximum of 26.5% of the dose 30 min after injection. Sephadex G-50 gel filtration profiles of radioactivity solubilized by Triton X-100 demonstrated that the radioactivity in kidney was mainly in the form of ¹³¹I-lysozyme. The intracellular distribution of TCA-precipitable ¹³¹I in the kidney was studied by the differential centrifugation of kidney homogenates during the first 3 hr after intravenous injection of ¹³¹I-lysozyme in rats. The specific radioactivity in the 19600×g (20 min) fraction of kidney was found to be the highest among another fractions. The proteinbound radioactivity detected in the 19600×g fraction of kidney 30 min after injection was suggested to be present in phagolysosomes by sucrose density gradient centrifugation and solubilization with Triton X-100. The degradation of injected ¹³¹I-lysozyme by the 19600×g fraction or phagolysosomes was examined, in succession. The pH-degradation profile of the 19600×g fraction was a monopeak with the optimum at pH 5.0. Based on the observation of the activation by cysteine and the inhibition by iodoacetamide, it was indicated that the proteolytic enzyme with the optimal pH of 5.0 in phagolysosomes is cathepsin B₁. The main degradation product of injected ¹³¹I-lysozyme in the phagolysosomes was identified as ¹³¹I-diiodotyrosine by paper chromatography. In contrast, the radioactivity excreted in urine was mainly inorganic ¹³¹I^-.