Intracellular coupling of bikunin and the heavy chain of rat pre-alpha-inhibitor in COS-1 cells.
Pre-alpha-inhibitor is a serum protein consisting of two polypeptides: bikunin of 16 kDa, which carries an 8 kDa chondroitin sulphate chain, and heavy chain 3 (H3) of 74 kDa. The two polypeptides are linked through an ester bond between an internal N-acetylgalactosamine residue of the chondroitin sulphate chain and the C-terminal aspartic acid residue of H3. Both bikunin and H3 are synthesized by hepatocytes and become linked as they pass through the Golgi complex. H3 is synthesized with both N- and C-terminal extensions which are released during intracellular transport. To be able to analyse the assembly of pre-alpha-inhibitor in detail, we have cloned and sequenced the cDNA of rat H3. Upon expression of the protein in COS-1 cells, both propeptides were found to be released. Furthermore, co-expression of H3 and bikunin resulted in the two polypeptides becoming coupled, indicating that cells other than hepatocytes may have the capacity to form chondroitin sulphate-containing links.
Blom
A M
AM
Department of Medical and Physiological Chemistry, Uppsala University, Biomedical Center, Sweden.
Thuveson
M
M
Fries
E
E
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D014162:Transfection; D014360:Trypsin Inhibitor, Kunitz Soybean; D014361:Trypsin Inhibitors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
D018076:DNA, Complementary; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D011498:Protein Precursors; C109148:SPINT2 protein, human; D014361:Trypsin Inhibitors; C061151:pre-alpha-trypsin inhibitor; D014360:Trypsin Inhibitor, Kunitz Soybean