The presence of discrete bands which differ in size

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The presence of discrete bands, which differ in size by several hundred amino acids, requires some additional explanation. If the zippering process were absolutely smooth, and all trypsin-cleavage sites were equally sensitive, A452 Chapter 19: Cell Junctions, Cell Adhesion, and the Extracellular Matrix
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then a whole series of bands would be expected, each differing from the last by an increment reflecting the spacing of the trypsin-sensitive bonds (lysines and arginines). The absence of this finely spaced ladder of bands suggests either (1) that the cleavage sites are not equally sensitive (the trypsin digestions were very brief in these experiments) or (2) that zippering is not smooth, but rather, pauses at characteristic points that represent more difficult regions to assemble. It is important to realize that the reassembly results tell you nothing directly about the natural assembly of type III collagen. It could be argued reasonably that the disulfide bonds at the C-terminus provide an artificial nucleation site for reassembly that is not present naturally. Nevertheless, these reassembly results are consistent with other in vivo observations. For example, point mutants of collagen typically are better assembled on the C-terminal side of the mutation than on the N-terminal side. This observation suggests that assembly is initiated toward the C-terminus and progresses zipperlike toward the N-terminus. A zipperlike assembly from one end also makes good theo- retical sense: in such a highly repetitious molecule as collagen, random inter- nal initiation would rarely lead to correctly aligned polypeptides. All these considerations, taken together, form the basis for the current belief that fibrillar collagens assemble zipperlike from the C-terminus and that a primary function of the C-terminal propeptide is to hold the polypep- tides in register for proper assembly. Reference: Bachinger HP, Bruckner P, Timpl R, Prockop DJ & Engel J (1980) Folding mechanism of the triple helix in type III collagen and type III pN- collagen. Eur. J. Biochem. 106, 619–632. 19–86 A. The sticking of cells to the dishes means opposite things in the two experi- ments. In the first experiment, cell sticking indicates that the peptide is active. In the second experiment, it means the peptide is inactive. In the first experiment, the peptides are stuck to the dish. Only when the peptides con- tain the active segment will the cells stick to the peptides and, hence, to the dish. In the second experiment, the cells will stick to the fibronectin on the dish unless the receptor sites on the cell surface are already occupied by the small peptide, in which case binding to the dish will be inhibited. The two experiments represent alternative ways of measuring the same thing, namely, the specific interaction between a receptor on the cell surface and a ligand in the cell’s environment.
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  • Spring '04
  • EricLander
  • cell biology, Collagen, basal lamina, cell adhesion, J. Cell Biol.

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