Chemistry of the collagen cross-links. Nature of the cross-links in the polymorphic forms of dermal collagen during development

Abstract

Both the type I and type III collagens present in embryonic dermis are stabilized by the intermolecular cross-link, hydroxylysino-5-oxonorleucine, derived from hydroxylysinealdehyde, although the type I collagen possesses a significant proportion of dehydrohydroxylysinonorleucine. However, concurrent with the change in the proportion of the two types of collagen during postnatal development there is a change-over with both type I and III collagens to the labile cross-link, dehydrohydroxylysinonorleucine, derived from lysine aldehyde. The results indicate that the change in the nature of the cross-link with development is determined primarily by the change in the extent of hydroxylation of the lysine residues in the terminal non-helical regions rather than being due to the change in the type of collagen. (Epstein, 1974). Our own studies (Bailey & Robins, 1972) have demonstrated a corresponding change-over in the nature of the cross-links duri the early growth period from a stable intermolecular cross-link, derived from hydroxyallysine, to a chemically labile cross-link, derived from allysine. Therefor-we decided to demonstrate whether type III collagen also possesses lysine-derived cross-links in addition to its intramolecular disulphide bonds, and if so, whether there is a change in type of cross-link during development.In the present paper we report that such a change does occur in the nature of -the intermolecular cross-links of both types of collagen during the early growth period, thus confirming the previous preliminary results Ltd., Poole, Dorset, U.K. MethodsSkins were obtained from a 3-month foetal calf and a 9-month-old steer and were scraped clean of adhering fat, muscular tissue and the epidermis. The tissue was minced, and then extracted with M-NaCI/0.05M-Tris/HCl, pH7.5, for 2 days at 4°C. Separation of collagen types. Pepsin digestion and separation of type I and type III collagens were carried out as described by . Briefly, the insoluble residue from the salt (1 M-NaCI, pH7.4, 4°C for 2 days) and acid (0.5M-acetic acid, 4°C for 2 days) extractions was digested with pepsin at a substrate/enzyme ratio of 10:1 (w/w) for 19h at 5YC. After inactivation of the pepsin by raising the pH, the solubilized collagen was purified by precipitation with 5% (w/v) NaCi at 20°C, redissolved in 1 M-NaCl at 40C, and then subjected to fractional precipitation to obtain type III collagen at 1.6M-NaCl and type I collagen at 2.5M-NaCl.Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The type I and type III collagens were redissolved, denatured in sodium dodecyl sulphate (2 %, w/v) at 38°C and analysed for subunit composition by polyacrylamide-gel electrophoresis. The electrophoresis was carried out by using a flat-bed technique with 6.5 % (w/v) polyacrylamide gel, 211

Ultrastructural and Biochemical Studies on a Case of Elastosis perforans serpiginosa

“…Ft is known from the literature that embryonic tissues have a higher activity in hydroxylating enzymes than adult tissues [23]. This explains the high proportion in reduced embryonic tissue of hydroxylysine-derived cross-links as DHLNL and HLNL [18], The extent of hydroxylation we measured in EPS dermis was very similar to that of newborn dermis, confirming the presence of immature collagen fibers in the disease.…”

“…[17] and Bailey and Sims [18]. For a direct comparison, normal and pathological cleaned dermis were reduced in the same experiment.…”

The location of three collagen types in skeletal muscle

“…The precipitates obtained were reprecipitated and shown to be a single type of collagen by SDS-polyacrylamide gel electrophoresis [6] .…”

“…Collagen Types I and III were prepared from 248 foetal calf skin fo~o~g pepsin digestion and fractional precipitation with sodium chloride [5,6] . The precipitates obtained were reprecipitated and shown to be a single type of collagen by SDS-polyacrylamide gel electrophoresis [6] .…”

Age-Related Changes in the Reducible Cross-Links of Human Dermis Collagen

“…These cross-links are labile aldimine bonds which can be detected upon reduction with tritiated borohydride, act as intermediary compounds and are converted during the maturation process into a nonreducible and more stable form [3][4][5][6].…”