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-α-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, α1-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 α1-microglobulin-part of the α1-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 α1-microglobulin-part is not necessary for the first step of galactosaminoglycan assembly.