Immunological identification of two proteoglycan fragments derived from neurocan, a brain-specific chondroitin sulfate proteoglycan.
Neurocan is a brain-unique chondroitin sulfate proteoglycan (CSPG) whose expression and proteolytic cleavage are developmentally regulated. One of the proteolytic products (C-terminal half) is known to be a CSPG with a 150 kDa core glycoprotein (CSPG-150). To identify the N-terminal half of neurocan, we raised an anti-neurocan polyclonal antibody (PAb 291) using a synthetic peptide whose amino acid sequence matched a part of the N-terminal half of neurocan. Western blots showed that PAb 291 recognized two CSPGs, one with a 220 kDa core glycoprotein (CSPG-220, namely neurocan) and one with a 130 kDa core glycoprotein (CSPG-130) isolated from young rat brains. CSPG-130 was co-purified along with CSPG-220 by PAb 291-immunoaffinity column chromatography. The amino acid sequence of the N-terminus of the immunopurified CSPG-130 was exactly the same as the N-terminal sequence of CSPG-220. These results suggest that not only the C-terminal half (CSPG-150) but also the N-terminal half (CSPG-130) of CSPG-220 exists in a CSPG form in rat brain. Using PAb 291 and monoclonal antibody 1G2 (MAb 1G2) which recognizes CSPG-150 in addition to CSPG-220, we found that the contents of CSPG-130 and CSPG-150 in the rat brain reached maximum levels around the time of birth. Both CSPG-130 and 150 were observed, while CSPG-220 was hardly detectable in extracts from the adult rat brain. Immunohistochemical investigation showed that the PAb 291 antigen had a similar distribution pattern to the MAb 1G2 antigen.(ABSTRACT TRUNCATED AT 250 WORDS)
Department of Perinatology, Institute for Developmental Research, Aichi, Japan.
F
F
Matsui
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Watanabe
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Oohira
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000196:embryology / Q000254:growth & development / Q000378:metabolism*; D011508:Chondroitin Sulfate Proteoglycans / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D007118:Immunoassay; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D037181:Lectins, C-Type; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000032:analysis*; D058580:Neurocan; D010446:Peptide Fragments / Q000032:analysis* / Q000302:isolation & purification; D011499:Protein Processing, Post-Translational; D051381:Rats; D017207:Rats, Sprague-Dawley
D011508:Chondroitin Sulfate Proteoglycans; D037181:Lectins, C-Type; D009419:Nerve Tissue Proteins; D058580:Neurocan; D010446:Peptide Fragments; C076817:NCAN protein, human