Date of Award
1991
Degree Type
Dissertation
Degree Name
Doctor of Philosophy
Abstract
Ferritin can be isolated from quail reticulocyte lysates using a rapid two-step purification method that combines heat denaturation and proteinase-K treatments to produce high yields of pure, undegraded ferritin. Ferritin remains in the supernatant of heat-denatured samples, and subsequent treatment with proteinase-K eliminates non-ferritin proteins that remain in the supernatant as well as non-ferritin proteins that are found in association with ferritin.;Ferritin is a heat shock protein of quail reticulocytes. Its synthesis in heat-shocked cells is 3-5 times greater than in control cells. Ferritin subunits synthesized in response to heat shock consist of isoforms that are identical in size and pI to those present in ferritin synthesized in control cells. The mechanism of this increased synthesis involves translational control.;The "prosome" fraction isolated from quail reticulocyte lysates by sucrose density gradient ultracentrifugation contains two biochemically and morphologically distinct particles. The major constituent ({dollar}\sim{dollar}95%) of this fraction is the proteasome, an M{dollar}\sb{lcub}\rm r{rcub}{dollar} {dollar}\sim{dollar}562,000 particle that possesses proteolytic activity. The proteasome is a cylinder {dollar}\sim{dollar}15 x 10 x 10 nm, and is composed of a set of protein subunits ranging in size from {dollar}\sim{dollar}22-{dollar}\sim{dollar}32 kDa. One of the subunits of the proteasome cross-reacts with a monoclonal antibody to prosome protein P27. The proteasome is composed of a spherical central component ({dollar}\sim{dollar}10 nm diameter) enclosed by two, 2.5 nm thick hexagonal end caps. The prosome particle is formed by degradation of the proteasome particle during sample preparation for electron microscopy. The minor constituent ({dollar}<{dollar}5%) of the prosome fraction is ferritin, a 12-13 nm diameter particle composed of 21 kDa subunits, which is found as the protein component of an M{dollar}\sb{lcub}\rm r{rcub}{dollar} {dollar}\sim{dollar}877,000 particle.;Ferritin is also the protein component of the {dollar}\sim{dollar}35S "prosome-like-particle", where it is found in association with small RNAs ({dollar}\sim{dollar}80 nucleotides in length) that hybridize to a globin cDNA.
Recommended Citation
Blaker, Timothy W., "Biochemical And Morphological Characterization Of Cytoplasmic Particles From Avian Reticulocytes" (1991). Digitized Theses. 2061.
https://ir.lib.uwo.ca/digitizedtheses/2061