In acute inflammation, the chondroitin-4 sulphate carried by bikunin is not only longer, it is also undersulphated

Abstract

Bikunin (Bk) is a Künitz-type serine proteinase inhibitor, which occurs in human plasma, mainly as covalent complexes with one or two of the three peptide heavy chains. The leading member of this glycoprotein family is inter-alpha-inhibitor (I alpha I), which consists of two heavy chains (H1 and H2) linked to Bk. Bk carries a glycosaminoglycan (GAG) chain, which is linked by ester bonds to the heavy chains of I alpha I. Furthermore, Bk, I alpha I and related components such as pre-alpha-inhibitor (P alpha I), all together making up the I alpha I family, present antiinflammatory and antimetastatic effects that hinge on this GAG chain. Recently (Eur. J. Biochem. 268 (2001) 2717), we provided evidence that, during acute phase response, the GAG chain of Bk, which is a low-sulphated chondroitin-sulphate, increases in size according to the severity of the inflammatory disease. This increase affects Bk-containing proteins in circulating blood as well as Bk excreted in higher amounts in urine of these patients. In this work, we have more extensively analysed the GAG chain of Bk isolated from urine collected from a unique patient with septic shock. Using MALDI-TOF-MS and HPLC analyses of chondrodisaccharides released by enzymatic digestion, we have demonstrated that the GAG chain is clearly modified; it consists of 20 +/- 5 disaccharide units vs. 14 +/- 3 for reference Bk originating from healthy donors. Among them, only 3 +/- 2.5 units are 4-sulphated for patient's Bk vs. 5 +/- 1.5 for reference Bk. Therefore, the non-sulphated region of the GAG chain, which is located towards its non-reducing end, where the heavy chains are positioned, is lengthened from 9 for reference Bk to 17 disaccharide units. We suggest that the biological effects of Bk-proteins may hereby be modulated during inflammatory diseases.