Expression of a functional proteinase inhibitor capable of accepting xylose: bikunin.
Bikunin is a Kunitz-type proteinase inhibitor, which is cross-linked to heavy chains via a chondroitin sulfate chain, forming inter-alpha-inhibitor and related molecules. Rat bikunin was produced by baculovirus-infected insect cells. The protein could be purified with a total yield of 20 mg/liter medium. Unlike naturally occuring bikunin the recombinant protein had no galactosaminoglycan chain. Endoglycosidase digestion also suggested that the recombinant form lacked N-linked oligosaccharides. Bikunin is translated as a part of a precursor, alpha1-microglobulin/bikunin, but the functional significance of the cotranslation is unknown. Our results indicate that the proteinase inhibitory function of bikunin is not regulated by the alpha1-microglobulin-part of the alpha1-microglobulin/bikunin precursor since recombinant bikunin had the same trypsin inhibitory activity as the recombinant precursor. Both free bikunin and the precursor were also functional as a substrate in an in vitro xylosylation system. This demonstrates that the alpha1-microglobulin-part is not necessary for the first step of galactosaminoglycan assembly.
Falkenberg
C
C
Department of Cell and Molecular Biology, Lund University, Sweden. cecilia.falkenberg@medkem.lu.se
Wester
L
L
Belting
M
M
Eklund
E
E
Akerström
B
B
D000818:Animals; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis*; D015842:Serine Proteinase Inhibitors / Q000096:biosynthesis* / Q000235:genetics; D014357:Trypsin / Q000187:drug effects; D014360:Trypsin Inhibitor, Kunitz Soybean; D014994:Xylose / Q000378:metabolism*
D008562:Membrane Glycoproteins; D011498:Protein Precursors; D011994:Recombinant Proteins; C109148:SPINT2 protein, human; D015842:Serine Proteinase Inhibitors; D014360:Trypsin Inhibitor, Kunitz Soybean; D014994:Xylose; D014357:Trypsin