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Title: Ribonucleases in rat liver
Author: Burge, Malcolm Leonard Ernest
ISNI:       0000 0001 3509 4973
Awarding Body: University of Surrey
Current Institution: University of Surrey
Date of Award: 1973
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Several precipitants commonly used in ribonuclease assays were compared with regard to the size of oligonucleotides remaining in solution. Centrifugation in HS and A XII zonal rotors was used to determine the distribution of rat liver ribonucleases among the various organelles. Experiments were performed to discover the best gradient and centrifugation conditions for the purification of lysosomes and mitochondria. Such experiments demonstrated the heterogeneity of lysosomes. Separation after pretreatment with Triton WR-1339 showed acid ribonuclease-rich lysosomes were more active in taking up the detergent than acid phosphatase-rich lysosomes. Isopycnic density gradient centrifugation of material from regions of the zonal run confirmed this observation and gave lysosomes purified about 20 times. The distribution of other lysosomal enzymes in fractionated normal and Triton-treated liver indicated further lysosomal heterogeneity. After fractionation of crude mitochondrial/ lysosomal fraction on an HS zonal rotor, alkaline ribonuclease showed a complex distribution. Comparison with that of marker enzymes suggested that the major part of the activity was due to lysosomes and plasma membrane. The lack of latency of the lysosomal enzyme suggested that it was mainly located on the lysosomal membrane. Fractionation of crude nuclear fraction on an A XII rotor showed that about half the particulate alkaline ribonuclease and most of the alkaline phosphodiesterase was in plasma membrane fragements. Alkaline RNase was also located in the mitochondria. Some studies were made on the properties of the plasma membrane, mitochondrial and lysosomal alkaline ribonucleases. Hepatoma samples, fractionated in both rotors, gave confusing results because of the reduced and extremely widely varying size of the mitochondria and lysosomes. 4'F-DAB-induced precancerous liver was investigated to determine whether the increased cytosol acid RNase resulted from lysosome breakdown. Treatment with actinomycin D induced an increase in total and cytosol acid RNase. The presence of acid and alkaline RNase activity in aqueous nuclei preparations was confirmed. Acid RNase was more active in nuclei from ethionine-fed rats.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available