| Description |
The structure and metabolism of elastin and collagen were studied in three sets of experiments. In the first experiment, porcine tropoelastin was digested with the endopeptidase, thermolysin. The digest contained at least 35 groups of peptides. The polydispersed peptides included large nonpolar peptides, 50-60 residues in length, and a series of gradations to polar peptides of 10 residues and less. The largest peptide sequenced was a portion of the tryptic peptide T1, which contained the pentapeptide repeat P G V G V. This region was enzyme resistant, implying that the peptide was in some sort of conformation unavailable to the thermolysin. Very likely, it is similar to the B-spiral which is composed of the pentapeptide repeat proposed by Urry. Two small polar peptides were also sequenced. One was shown to extend 2 residues amino terminal to peptide T7a. The other was previously unknown, its sequence being: I G G K P P K P G. Because of the two lysines and the rigid proline structure, it may be involved in some sort of crosslinking function. Overall, thermolysin appeared to cleave the hydrophilic crosslink regions of tropoelastin more extensively than the large hydrophobic regions. In the second experiment, elastin and collagen metabolism were analyzed from the upper thorax to the abdomen in the young pig aorta, in vitro. Tissue was incubated with either ('14)C leucine, ('14)C valine, ('3)H valine, or ('3)H proline for 1, 2, or 24 hours. The tissue was extracted with detergent, urea, and autoclaving. Total incorporated radioactivity was higher in the upper thorax than in the abdomen during the 1 or 2 hour incubations. Most of the radioactivity was seen in valine-rich soluble proteins. However, after a 24 hour incubation the abdomen was the most radioactive region. The label was primarily in the autoclaved supernate, and it reflected collagen formation as shown by ('3)H hydroxyproline production. From this, it was concluded that collagen formation did not appear to be altered significantly in vitro, while elastin formation was markedly altered. Thus, quantitative questions about elastin metabolism must be answered through experiments conducted in vivo. The third experiment involved analyzing elastin and collagen metabolism from the upper thorax to the abdomen of 4-6 week and 2 day old rat aortas, in vivo. Rats were injected intraperitoneally with ('3)H glycine and then sacrificed at specified times after injection. Soluble and insoluble elastin and collagen were isolated. In the 2 day old rat little radioactivity in tropoelastin was isolated. The elastin showed an increase in the accumulation of radioactivity in the upper thorax as compared to the abdomen. Synthesis of radioactive soluble collagen was higher in the abdomen, but this was not reflected by increased accumulation of radioactivity into insoluble collagen. Finally, radioactivity in both elastin and collagen appeared to accumulate at the same relative rates. In the 6 week old rat, again little radioactivity in tropoelastin was isolated. This indicated that tropoelastin was rapidly crosslinked or otherwise utilized. Accumulation of radioactivity in elastin of the upper thorax was greater than in the abdomen. Synthesis of radioactive collagen was increased in the abdomen along with increased accumulation of radioactivity into insoluble collagen. Finally, accumulation of radioactivity in collagen appeared to proceed slower than in elastin |
