Interaction between Membrane Functions and Protein Synthesis in Reticulocytes
- 1 September 1978
- journal article
- research article
- Published by Wiley in European Journal of Biochemistry
- Vol. 89 (2), 341-352
- https://doi.org/10.1111/j.1432-1033.1978.tb12535.x
Abstract
An inhibitor of protein synthesis was isolated from reticulocyte membranes by solubilization with Triton X-100 and purified using heat treatment, filtration on Amicon filters, DEAE-cellulose ion-exchange chromatography and Sephadex G-75 gel chromatography. A final purification of 120-fold was achieved. The purified inhibitor was 95% homogenous when run on a dodecylsulfate/polyacrylamide gel system. Three independent methods were used to estimate the MW of the purified inhibitor: Sephadex G-75 gel chromatography, dodecylsulfate/polyacrylamide gel electrophoresis and sucrose gradient all confirmed that the purified inhibitor was a small molecule with a sedimentation coefficient of 0.7 S and a MW ranging 5000-8000. The purified inhibitor possesses a specific endonucleolytic activity, degrading the 28-S species of rRNA to species sedimenting between 10-14 S. Due to its membrane localization the name RNase M is proposed. The purified inhibitor''s endonucleolytic activity was characterized with regard to its kinetics, concentration dependence, pH optimum and its requirements for divalent cations. Kinetics showed that RNase M retained its specificity after 60 min of incubation with the RNA substrate. Specificity was also demonstrated by incubating the polysomal RNA with high concentrations of purified enzyme. The pH optimum was between pH 6 and pH 7, and the enzyme did not require divalent cations for its activity. Pancreatic RNase B used at a similar protein synthesis inhibitory concentration as the RNase M caused a complete breakdown of rRNA to oligonucleotides and mononucleotides. The possible biological significance of the purified inhibitor in regulating protein synthesis in the maturing reticulocyte is discussed.This publication has 22 references indexed in Scilit:
- Interaction between Membrane Functions and Protein Synthesis in Reticulocytes: Specific Cleavage of 28‐S Ribosomal RNA by a Membrane ConstituentEuropean Journal of Biochemistry, 1978
- Specific degradation of ribosomal RNA in rabbit reticulocyte membrane‐bound ribosomesFEBS Letters, 1977
- Intracellular distribution of ribonuclease activity during erythroid cell developmentBiochimica et Biophysica Acta (BBA) - Nucleic Acids and Protein Synthesis, 1977
- Phosphorylation of initiation factor eIF-2 and the control of reticulocyte protein synthesisCell, 1977
- Mechanism of Action of Colicin E3. Effect on Ribosomal Elongation-Factor-Dependent ReactionsEuropean Journal of Biochemistry, 1977
- Interaction between Membrane Functions and Protein Synthesis in Reticulocytes. A Membrane Constituent which Inhibits Protein Synthesis in Cell-Free SystemEuropean Journal of Biochemistry, 1976
- Partial purification of a translational repressor mediating hemin control of globin synthesis and implication of results on the site of inhibitionBiochemical and Biophysical Research Communications, 1973
- Erythroid cell RNase: activation by urea and localization to the cell membraneJCI Insight, 1971
- Effect of ribonuclease on Escherichiacoli ribosomesBiochemical and Biophysical Research Communications, 1970
- Characteristics of RNA degradation in the erythroid cellJCI Insight, 1969