Tropoelastin protein monomers associate by coacervation and are cross-linked in vivo to form elastin macro-assemblies. We provide evidence for specific protein domain contact points between tropoelastin monomers during association by coacervation. The homobifunctional cross-linker bis(sulfosuccinimidyl) suberate served as a rapid reporter of adjacent lysines and preferentially exposed domains. Intact cross-linked peptide pairs were identified after protease digestion and high-resolution electrospray mass spectrometry followed by MS/MS sequencing. Mapping of the assigned sequences indicated that the region in the monomer spanning domains 19-25 was readily accessible to solvent and enriched in cross-linking. Domains 12 and 36 were also prevalent, Where these two regions were not previously thought to play a major role ill the formation of mature elastin. A specificity for particular lysines allowed for the construction of a model for the first close contacts between domains and the first detailed study of the cross-linking of tropoelastin. (C) 2004 Elsevier Inc. All rights reserved.