. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. COLLAGEN IN GORGONIAN SPICULES B Figure 3. SDS-PAGE of the traction of the insoluble matrix not soluhilized in acetic acid and pepsin, stained for (A) protein with Com- massie blue, and (B) carbohydrate with PAS. above (Bonar and Glimcher, 1970). The physiological, relatively gentle digestion procedure used in this study degraded and solubilized all the collagenous and non-col- lagenous protein that was not protected by mineral. Pre- vious methods used to isolate "calcined" structures used NaOCl or a strong base to
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. COLLAGEN IN GORGONIAN SPICULES B Figure 3. SDS-PAGE of the traction of the insoluble matrix not soluhilized in acetic acid and pepsin, stained for (A) protein with Com- massie blue, and (B) carbohydrate with PAS. above (Bonar and Glimcher, 1970). The physiological, relatively gentle digestion procedure used in this study degraded and solubilized all the collagenous and non-col- lagenous protein that was not protected by mineral. Pre- vious methods used to isolate "calcined" structures used NaOCl or a strong base to destroy organic material. How- ever, NaOCl will produce anorganic bone from normal mammalian bone and is often used to examine bone ar- chitecture by scanning electron microscopy. This also de- grades as well as destroys a significant portion of the or- ganic matrix of mineralized tissue. The above analysis is substantiated by a comparison of the current results with a previous failure to detect collagen in the insoluble matrix fraction (Kingsley and Watabe, 1983). Kingsley and Wa- tabe (1983) isolated spicules in NaOCl and demin- eralized in N HC1 at room temperature. The isolated spicules that were analyzed were, in both cases, from an- imals collected during the summer, so the previous meth- ods were harsh and yielded erroneous results. The spicule insoluble matrix of the gorgonian Briareitm asbestinum consisted of very small peptides (1600-5000 daltons) and contained 22 Hyp residues/1000, approxi- mately 20% Gly and no Hyl (Silberberg el ai. 1972). These features only suggest the presence of collagen-like com- ponents. The amino acid composition of the whole spicule matrix of the gorgonian Pseudoplexaura flagellosa re- vealed 48, 237, and 9 residues/1000 total residues of Hyp, Gly, and Hyl, respectively (Goldberg, 1988). Silberberg el ai (1972) air dried, ground the whole animals, digested the organic material with 30% KOH, and demineralized in concentrated HC1 a
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