Proteins of crustacean exoskeleton: IV. Partial amino acid sequences of exoskeletal proteins from the Bermuda land crab, Gecarcinus lateralis, and comparisons to certain insect proteins.

Abstract

As in all decapod Crustacea, the exoskeleton of the land crab Gecarcinus lateralis consists of four layers. Prior electrophoretic analysis of proteins extracted from these layers revealed an abundance of small M(r) proteins with acidic pIs are found in insect cuticle (O'Brien et al. [1991 Biol. Bull., 181:427-441). Further, immunological cross-reactivity between crab exoskeletal proteins and insect cuticular proteins has been demonstrated (Kumari and Skinner [1993] J. Exp. Zool., 265:195-210). Partial amino acid sequences of a number of proteins from the four exoskeletal layers are described here. Proteins were electrophoresed on two-dimensional (2D) gels, transferred to polyvinylidene difluoride (PVDF) membranes, and stained; individual spots were recovered and their N-termini were sequenced. In addition, a 14-kDa protein (pI = 5.4) from membranous layer (ML14) was eluted from 2D gels and digested with endoproteinase Lys-C; N-termini of its constituent peptides were sequenced. The two epicuticular proteins differed from each other. Three proteins with identical electrophoretic mobility isolated from exocuticle, endocuticle, and membranous layer appeared to have identical N termini, while another electrophoretically identical set from the three layers appeared identical with each other but differed in three positions from the first set. Two proteins from the membranous layer both had a mass of 25 kDa but different isoelectric points. Their sequences were indistinguishable from each other but clearly distinct from another membranous layer protein. Another distinct sequence was found in a 14-kDa protein from endocuticle, while a less acidic pair of 14-kDa proteins from endocuticle and membranous layer were quite similar to one another. The three internal peptide fragments from ML14 were distinct, but one had regions similar to the ML14 N terminus. One crab exoskeletal protein sequence was similar to some structural proteins of vertebrates, whereas others had motifs found in insect cuticular proteins. The sequence similarities identified did not account for the antibody cross-reactivity.