Type IX collagen is a recently described component of cartilage. This nonfibril-forming molecule is composed of three helical (COL1-COL3) and four nonhelical domains (NC1-NC4) and also serves as a core protein for a single glycosaminoglycan side chain. To understand its spatial relationship with other matrix constituents, a peptide containing a lysine-derived cross-link was isolated and subjected to amino acid sequencing. The results show that type IX collagen is linked to type II collagen by a hydroxypyridinium cross-link. This cross-link connects the central triple helical (COL2) domain of one alpha 2(IX) chain to the amino telopeptides of two alpha 1(II) chains. The cross-link is very close to the glycosaminoglycan attachment site found in the nonhelical NC3 domain of the alpha 2(IX) chain. A model of the resulting assembly suggests that type IX collagen is located at the surface of the fibril with the short COL 3 domain and the basic globular NC4 domain projecting out from the surface of the fibril. The NC4 domain could, therefore, serve as a binding site for other matrix constituents. In this model, the glycosaminoglycan chain of type IX collagen would be located at the gap region of the fibril.