Zinc metalloprotease activity in the cement precursor secretion of the barnacle, Chthamalus fragilis Darwin

• Published in: Tissue & cell Vol. 28; no. 4; pp. 439 - 447
• Main Author: Dougherty, W.J.
• Format: Journal Article
• Language: English
• Published: Scotland Elsevier Ltd 01.08.1996
• Subjects: Barnacle; cement; Chthamalus fragilis; Marine; secretion; Zn-metalloprotease; Barnacle; Zn-metalloprotease; cement; secretion
• ISSN: 0040-8166; 1532-3072
• DOI: 10.1016/S0040-8166(96)80029-2

Adult barnacles, Chathamalus fragilis, were removed carefully from the leaves and stems of marsh grass and floated base-up in algae-supplemented sea water. During the next 24–72 h, the animals secreted onto their exposed bases, a fluid, the cement precursor secretion (CPS), aliquots of which were collected in glass micropipets and pooled. The concentration of protein in pooled samples of CPS averaged 1.5 g ± 0.42 protein/l of secretion. Protease activity was expressed as A 492 units/h/g of CPS protein. Aliquots of 5–45 l of pooled CPS samples, incubated in the presence of FTC-casein at 37°C for 24 h, exhibited 8.31 × 10 −5 ± 1.55 × 10 −5 ΔA 492 units/hr/g of protein on average. Protease activity was optimized by the addition of 10 mM Ca ++ ions. Activity was detectable over a broad pH range, but was optimal around pH 8. Protease activity was inhibited up to 40% in the presence of 2.7 mM ethylenediaminetetraacetic acid (EDTA) and up to 97% in the presence of 2.5 mM 1,10-phenanthroline (OP) in the presence of 10 mM Ca ++ ions. Although low concentrations of Zn ++ ions (10 M) had little effect on protease activity, higher concentrations of Zn ++ ions (50 M to 15 mM Zn ++) inhibited CPS protease activity. Protease activity was not inhibited by 1 mM phenylmethylsulfonylfluoride (PMSF), nor by 28 M E64, nor by 20 M leupeptin. At the present time, the protease activity in the barnacle CPS may best be characterized as a Ca-stimulated Zn-metalloprotease.