Type XIII collagen forms homotrimers with three triple helical collagenousdomains and its association into disulfide-bonded trimers is enhanced by prolyl 4-hydroxylase

Type XIII collagen is a type II transmembrane protein predicted to consist of a short cytosolic domain, a single transmembrane domain, and three colla genous domains flanked by noncollagenous sequences. Previous studies on mRN As indicate that the structures of the collagenous domain closest to the ce ll membrane, COL1, the adjacent noncollagenous domain, NC2, and the C-termi nal domains COL3 and NC4 are subject to alternative splicing. In order to e xtend studies of type XIII collagen from cDNAs to the protein level we have produced it in insect cells by means of baculoviruses. Type XIII collagen alpha chains mere found to associate into disulfide-bonded trimers, and hyd roxylation of proline residues dramatically enhanced this association. This protein contains altogether eight cysteine residues, and interchain disulf ide bonds could be located in the NC1 domain and possibly at the junction o f COL1 and NC2, while the two cysteine residues in NC4 are likely to form i ntrachain bonds. Pepsin and trypsin/chymotrypsin digestions indicated that the type XIII collagen alpha chains form homotrimers whose three collagenou s domains are in triple helical conformation. The thermal stabilities (T-m) of the COL1, COL2, and COL3 domains were 38, 49 and 40 degrees C, respecti vely. The T-m of the central collagenous domain is unusually high, which in the light of this domain being invariant in terms of alternative splicing suggests that the central portion of the molecule may have an important rol e in the stability of the molecule. All in all, most of the type XIII colla gen ectodomain appears to be present in triple helical conformation, which is in clear contrast to the short or highly interrupted triple helical doma ins of the other known collagenous transmembrane proteins.