Properties of beef pancreas ribosomes.
Creators: name: Madison, James Thomas
Publisher: name: University of Utah
Year: 2012
Subjects: Cell Fracationation
Homogenate

description: A. Ribosomes have been isolated from beef pancreas microsomes and compared to similar particles prepared from rat liver.  B. The composition of the two types of ribosomes was very similar, except that the beef pancreas particles contained about 1,000 times more ribonuclease activity.  The pancreas particles contained about 2% chymotrypsinogen and 1% ribonuclease.  C. Sperimine (0.1 mM) and potassium chloride (0.1 M) released a large portion of the beef pancreas ribosomal ribonuclease, indicating that the enzyme was bound by electrostatic forces.  D. About 25% of the beef pancreas ribosomal protein was soluble in ethanol if the particles had been precipitated with trichloracetic acid.  E. The ultracentrifugal patterns of beef pancreas and rat liver ribosomes were very similar.  Both types had a major component with a sedimentation coefficient of about 80 S.  The observed sedimentation coefficient of the pancreas ribosomes was strongly dependent on the concentration of the ribosomal suspension being analyzed.  F. Pancreases ribosomes did not require the addition of magnesium to maintain them in the 80 S form.  Overnight dialysis of well-washed pancreas particles against a magnesium-free medium resulted in the appearance of two additional peaks, in the ultracentrifuge, with sedimentation coefficients of about 115 S and 140 S.  Deoxycholate, but not ethylenediaminetetraacetate, produced subunits of 50 S and 30 S from beef liver pancreas ribosomes.  These observations suggest that the bonds between the pancreas subunits are stronger than the union between the liver subunits.  G. Incubation of beef pancreas ribosomes at room temperature, or above, resulted in an increase of A260 of approximately 25%.  At 26°C the hyper-chromic effect was usually complete in 60 to 90 minutes, while at 37°C the maximum A260 was reached in 10 to 15 minutes.  Incubation at 26°C also caused the beef pancreas ribosomes to aggregate.  When the increase in A260was complete, no 80 S particles remained.  H. Urea, ethylenediaminetetraacetate, and ionic strengths greater or less than 0.0003 accelerated the appearance of the hyper-chromic effect at 26°C.  I.  The addition of anti-ribonuclease serum, magnesium, calcium, spermine or sulfated polysaccharides, but not trypsin inhibitor or anit-chymotrypsinogen, prevented the hyper-chromic effect at 26°C suggesting that an attack by ribonuclease was required to initiate the events that led to the hyper-chromic effect.  J. Rat liver ribosomes exhibited no hyper-chromic effect at 26°C or 37°C in mM phosphate, pH 7.8.  The addition of ribonuclease (1 umg/ml.) or potassium chloride (0.1 M), however, resulted in an increase in their A260 upon incubation.  K.  Beef pancreas ribosomes, through very unstable at 26°C or 37°C, were found to be almost completely resistant to hydrolysis by ribonuclease at 4°C.  This result suggested that the RNA of the intact ribosome was not susceptible to attack by ribonuclease, but that small changes in the configuration of the particle would allow ribonuclease to act.  Freezing and thawing, on the other hand, allowed the RNA to dissociate from the protein of the pancreas ribosomes.  L.  Beef pancreas ribosomes were precipitated by mM magnesium or calcium chloride, while 10 mM magnesium chloride was required to precipitate rat liver ribosomes.  Pancreas ribosomes also had a lower anodic mobility upon electrophoresis than the rat liver particles.  These results indicate that basic proteins were bound on the surfaces of pancreas ribosomes.  M. Similarities and differences in the properties of beef pancreas and rat liver ribosomes have been summarized by the use of model structures.  It is proposed that binding of basic proteins by the beef pancreas particles during isolation seriously affected their properties.