Structural and Electron Microscopic Analysis of Neurocan and Recombinant Neurocan Fragments

Abstract

Neurocan, a nervous tissue-specific chondroitin sulfate proteoglycan of the aggrecan family which has been shown to interact with neural cell adhesion molecules and tenascin, could be visualized by rotary shadowing electron microscopy as two globular domains interconnected by an extended flexible filament of 60 -90 nm. Several recombinant neurocan fragments generated in the human embryonic kidney cell line 293 represent as observed by electron microscopy the expected parts of this structure, which indicates a correct folding of these molecules. Biological activity of the recombinant N-terminal globular domain could be demonstrated by its coelution with hyaluronan in gel permeation chromatography. In addition, the modification of the recombinant fragments with certain carbohydrate structures was analyzed. High mannose oligosaccharides could be mapped to the N-terminal globular domain of the brainderived molecule. Only recombinant fragments containing parts of the central region of the molecule were modified with chondroitin sulfate chains and with the HNK-1 epitope, and could be considerably altered in their migratory behavior on SDS-polyacrylamide gel electrophoresis by neuraminidase treatment. These findings and the electron microscopic shape indicate a mucin-like character for the central neurocan region.

MATRIX PROTEOGLYCANS: From Molecular Design to Cellular Function

“…NEUROCAN The third member of the hyalectan gene family is neurocan, a chondroitin sulfate proteoglycan originally cloned from rat brain (146). Rotary shadowing electron microscopy of tissue-derived neurocan reveals two globular domains connected by a central rod of 60-90 nm (147) Recombinant studies have demonstrated that glycanation is restricted solely to this central domain even though other portions of the molecule contain SG repeats (147). The C-terminal domain is again very similar to that of the other members of the hyalectan gene family with approximately 60% identity between the rat neurocan and human versican and aggrecan.…”

The interaction of versican with its binding partners

“…The characteristic features of hyalectans, also termed lecticans, are their similar globular structures at both N-and C-termini [4,16], and their lectin-like activity [17]. This sub-group of proteoglycans contains multiple domains and mediates a variety of cell activities [10,[18][19][20][21][22].…”

Extracellular matrix of the central nervous system: from neglect to challenge

“…1). Shared features among these large chondroitin sulfate proteoglycans are the highly homologous G1 and G3 domains, which appear in rotary shadowing electron microscopy as compact globular structures at either end of an extended, but Xexible central region (Mörgelin et al 1989;Retzler et al 1996). This poorly sequence-conserved middle part carries most of the O-and N-linked oligosaccharides and all glycosaminoglycan side chains.…”