Coacervation Is Promoted by Molecular Interactions between the PF2 Segment of Fibrillin-1 and the Domain 4 Region of Tropoelastin

Abstract

In forming elastic fibers, microfibrils act as the scaffold sites for depositing the elastin precursor tropoelastin. We examined key binding interactions that promote massive tropoelastin association through coacervation. Using a segment of the microfibril protein fibrillin-1, PF2, known to bind full-length tropoelastin, we mapped its interaction site to the N-terminal region of tropoelastin bounded by domains 2 and 18. Precise contact residues between domain 4 of tropoelastin and domain 16 of fibrillin-1 were discovered through a novel combination of transglutaminase cross-linking and mass spectroscopy, with contact sites at residues K38 of tropoelastin and Q669 of fibrillin-1. This is the first report of a role for this region of tropoelastin in microfibril interactions. The addition of PF2 thermodynamically facilitated the coacervation of tropoelastin, resulting in smaller changes in entropy and enthalpy values for the coacervating system. A novel multicomponent in vitro tropoelastin assembly reaction system demonstrated that amassed tropoelastin was spatially and preferentially directed to surfaces coated with PF2 as expected for organized three-dimensional distribution during tissue elastogenesis. This study underscores the role of this part of fibrillin-1 as an anchor point for tropoelastin at the microfibril-elastin junction during the initial stages of elastic fiber assembly.

Functional Consequences of Homocysteinylation of the Elastic Fiber Proteins Fibrillin-1 and Tropoelastin

“…The self-assembly process for tropoelastin and the properties of elastin-like polypeptides can be experimentally assessed in a temperature-induced phase separation, known as coacervation (25,26). This process is dependent on factors such as ionic strength, pH, polypeptide concentration, and the presence of other extracellular matrix proteins (27)(28)(29)(30)(31).…”

Microfibril-associated MAGP-2 Stimulates Elastic Fiber Assembly

“…Mutation of fibrillin-1 in the Tight-skin (Tsk) mice induces major alterations in microfibril structures (12,13). They also have a function in tropoelastin deposition as they regulate tropoelastin coacervation (14). They are large glycoproteins that have distinct but overlapping pattern of expression (15).…”

Tissue Engineering and Regenerative Strategies to Replicate Biocomplexity of Vascular Elastic Matrix Assembly

“…The tropoelastin is thus able to congregate on the microfibrillar scaffold as amorphous elastin and then undergo crosslinking. 9 The microfibrils are composed of several different glycoproteins, most prominently fibrillin, and serve to maintain the integrity of elastic fibers and enable VSMCs to interact with these structures. Crosslinking of elastin is initiated by oxidation of lysine amino acid side chains on the elastin molecules by a Cu 2 + -dependent enzyme, LOX.…”