Dog pancreas

By using an HeLa cell-free system together with microsomes from dog pancreas, four proteins can be made from the 26 S RNA the capsid protein, the E1 protein, a protein with an of 62,000 (the p62 protein),... 

Minkowski tried unsuccessfully to prepare an extract of dog pancreas that would reverse the effect of removing the pancreas—that is, would lower the urinary or blood glucose levels. We now know that insulin is a protein, and that the pancreas is very rich in proteases (trypsin and chymotrypsin), normally released directly into the small intestine to aid in digestion. These proteases doubtless degraded the insulin in the pancreatic extracts in Minkowski s experiments. 

By the turn of the twentieth century, it was realized that diabetes was associated with the pancreas. In 1921, Banting and Best successfully isolated insulin from dog pancreas, and this was followed quickly by tests in humans. Eli Lilly began production of insulin in 1923. 

Fig. 3. Removal of PDI from dog pancreas microsomes. Dog pancreas microsomes after the various treatments were isolated by ultracentrifugation and separated on a 129c polyacrylamide gel in the presence of SDS. The gel was loaded as follows lane I, molecular-weight markers lane 2, untreated microsomes lane 3, pH 9-washed microsomes lane 4, saponin-washed (1%, w/v) microsomes lane 5, sonicated microsomes lane 6, pH 9-washed microsomes reconstituted with purified PDI.

Fig. 5. Cell-free translation of -y-gliadin mRNA. The -y-gliadin niRNA was translated in the absence (lanes 1 and 5) or presence (lanes 2-4 and 6) of dog pancreas inicrosomes. Translation products were separated by SDS-PAGE under reducing conditions (lanes I -3, 5, and 6) and under nonreducing conditions (lane 4). Lanes 1 and 5 are products of translation in the absence of microsomal vesicles lanes 3,4, and 6 are products of translation in the presence of microsomal vesicles translation products of lanes 3 and 4 were treated with proteinase K translation products of lane 6 were treated with 1 (v/v) Friton X-100 and proteinase K.

Fig. 10. Cell-free synthesis of t-PA glycoforms. The niRNA coding for t-PA was translated in a rabbit reticulocyte lysate in the presence of dog pancreas microsomes. Microsonies were isolated posttranslationally and the translocated, glycosylated products were separated by SDS-PAGE. Translation was carried out under conditions that either prevented (lane 2) or allowed (lane 3) proper folding of the t-PA molecule, yielding enzymatically active protein that was sensitive to natural inhibitors and stimulators.

One of Ihe major triumphs of the 20th century occurred in 1922, when Banting and Best extracted insulin from dog pancreas. Advances in the biochemistry of insulin have been reviewed with emphasis on proinsulin biosynthesis, conversion of proinsulin lo insulin, insulin secretion, insulin receptors, metabolism, effects by sulfonylureas. and so on. ... 

Watanabe, S., et al. "Release of Secretin of Liquorice Extract in Dogs." Pancreas 1(5) 449454, 1986. Abstract. Sage... 

After tolbutamide administration, the granules in the pancreas disappear, the amount of extractable insulin is decreased. These effects have been detected when fragments of dog pancreas are incubated with tolbutamide in vitro. 

The first isolation of an active OST complex, composed of three subunits (ribo-phorin I and II, and OST48) succeeded from dog pancreas cells [17]. The ribo-phorins had been characterized earlier as possible receptors for ribosomes and thought to be involved in protein translocation [119-121]. Since OST activity could be depleted from detergent solubilized extract by anti-ribophorin I antibodies, it became clear that ribophorins are indeed constituents of OST [17], They are no longer believed to be involved in ribosome binding, but in accord with the co-translational nature of A-glycosylation, the identification of ribophorins as part of the OST may indicate a close location of the enzyme to the protein translocation channel. 

Yeast Mol. wt. (k) Essential Dog pancreas Hen oviduct Chieken liver Pig liver Human Human liver lymphocytes ... 

In Table 1 a compilation of OST complexes from different organisms and the homology between the various subunits is depicted. In most cases the genes have been cloned or partial sequence information is available to allow their classification. Compared to yeast, the maimnalian enzymes seem to be composed of fewer subunits. In the light of the discussion above, one may predict that additional subunits may be detected that have escaped detection so far. In fact, it was subsequently found that the trimeric OST complex from dog pancreas [17], contains in addition DADl (defender of apoptotic death) [18] which is 40% identical to Ost2p from yeast. The isolation of incomplete , but active complexes indicates that the respective subunits may be sufficient to constitute a catalytical core unit. 

Figure 3. Predicted membrane topology of the oligosaccharyltransferase subunits from yeast. The topology is based partially on direct or indirect experimental evidence (N-terminal sequencing, protease accessibility, glycosylation sites, HIS4 fusions) as discussed. The presentation is not to scale. For comparison, the corresponding homologues of the OST complex from dog pancreas are depicted which have the same predicted topology (not shown).

Wagle, S. R. Studies on biosynthesis of insulin by pH-5 enzymes-microsome system from fetal dog pancreas. Biochim. et Biophys. Acta 95, 180 (1965). 

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