Elastin coacervation

D.W. Urry R. Henze, P. Redington, M.M. Long, and K.U. Prasad, Temperature dependence of dielectric relaxations in a-elastin coacervate evidence for a peptide librational mode. Biochem Biophys Res Commun 128,1000-1006,1985a. 

Coacervation occurs in tropoelastin solutions and is a precursor event in the assembly of elastin nanofibrils [42]. This phenomenon is thought to be mainly due to the interaction between hydro-phobic domains of tropoelastin. In scanning electron microscopy (SEM) picmres, nanofibril stmc-tures are visible in coacervate solutions of elastin-based peptides [37,43]. Indeed, Wright et al. [44] describe the self-association characteristics of multidomain proteins containing near-identical peptide repeat motifs. They suggest that this form of self-assembly occurs via specific intermolecular association, based on the repetition of identical or near-identical amino acid sequences. This specificity is consistent with the principle that ordered molecular assembhes are usually more stable than disordered ones, and with the idea that native-like interactions may be generally more favorable than nonnative ones in protein aggregates. 

This coacervation process forms the basis for the self-assembly, which takes place prior to the crosslinking. The assembly of tropoelastin is based on an ordering process, in which the polypeptides are converted from a state with little order to a more structured conformation [8]. The insoluble elastic fiber is formed via the enzymatic crosslinking of tropoelastin (described in Sect. 2.1). Various models have been proposed to explain the mechanism of elasticity of the elastin fibers. 

Elastin-mimetic protein polymers have been fabricated into elastic networks primarily via y-radiation-induced, radical crosslinking of the material in the coacervate state [10]. Although effective, this method cannot produce polymers gels of defined molecular architecture, i.e., specific crosslink position and density, due to the lack of chemoselectivity in radical reactions. In addition, the ionizing radiation employed in this technique can cause material damage, and the reproducibility of specimen preparations may vary between different batches of material. In contrast, the e-amino groups of the lysine residues in polymers based on Lys-25 can be chemically crosslinked under controllable conditions into synthetic protein networks (vide infra). Elastic networks based on Lys-25 should contain crosslinks at well-defined position and density, determined by the sequence of the repeat, in the limit of complete substitution of the amino groups. 

Starcher, B. G, Saccomari, G., Urry, D. W. Coacervation and ion-binding studies on aortic elastin. Biochim. Biophys. Acta 310, 481 (1973)... 

The process of coacervation is finely tuned to the physiological conditions of the extracellular matrix. Optimal coacervation of human tropo-elastin occurs at 37 °G, 150 mM NaCl, and pH 7-8 (Vrhovski et al, 1997). The arrangement of sequences in tropoelastin is critical to this process of coacervation, where association through hydrophobic domains depends on their contextual location in the molecule (Toonkool et al., 2001b). Tropoelastin association rapidly proceeds through a monomer to tuner transition, with little evidence of intermediate forms (Toonkool et al, 2001a). 

Ito, S., Ishimaru, S., and Wilson, S. E. (1998). Effect of coacervated alpha-elastin on proliferation of vascular smooth muscle and endothelial cells. Angiology 49, 289-297. Jacob, M. P., Badier-Commander, C., Fontaine, V., Benazzoug, Y., Feldman, L., and Michel, J. B. (2001). Extracellular matrix remodeling in the vascular wall. Pathol. Biol. 49, 326-332. 

Jensen, S. A., Vrhovski, B., and Weiss, A. S. (2000). Domain 26 of tropoelastin plays a dominant role in association by coacervation./. Biol. Chem. 275, 28449-28454. Kagan, H. M., and Sullivan, K. A. (1982). Lysyl oxidase Preparation and role in elastin biosynthesis. Methods Enzymol. 82, 637-650. 

Urry, D. W., Starcher, B., and Partridge, S. M. (1969). Coacervation of solubilized elastin effects a notable conformational change. Nature 222, 795-796. 

Partridge et al. (W ib) observed that when elastin from ligamentum nuchae of cattle was repeatedly extracted with 0.2. 5 M oxalic acid at 100°C the fibers completely dissolved after about 5 hr total extraction. On dialysis in cellophane only about. 5 % of the total nitrogen of the reaction mixture diffused through the membrane. The bulk of the product was a protein which was soluble in distilled water or buffer solutions at temperatures below 2f>°C, but on raising the temperature of the solution in dilute buffer at pH 4-6 a coacervate phase consisting of liquid droplets separated. The soluble material was thus similar in properties to the hemi-elastin of Horbaczewski (1882). 

Partial hydrolysis of elastin by reagents other than organic acids also gives rise to a mixture of soluble proteins similar to a- and /3-elastin. Thus Wood (quoted by Hall et al, 1952) first demonstrated that partial hydrolysis with dilute sodium hydroxide yields a protein which forms a reversible coacervate on raising the temperature of its solutions. Later Wood (1958) showed that on prolonged heating in aqueous solution a-elastin is converted into an insoluble gellike form. Reconstituted fibers of heat-treated a-elastin resembled fibrous elastin in their elastic behavior and X-ray-dif-fraction pattern but imlike purified elastin they were dissolved by 1 % acetic acid at 100°C and by crystalline trypsin. 

Ioffe and Sorokin (1954) investigated a novel procedure for the hydrolysis of elastin using copper sulfate and 0.4 N barium hydroxide at 37°C for 60 hr. The first product of hydrolysis was a protein which resembled a-elastin in that it showed reversible coacervation on raising the temperature. This substance was subsequently degraded further to yield a soluble fraction and a fraction containing peptides. Alkaline hydrolysis was much more rapid in the presence than in the absence of copper ion. 

S. Ito, S. Ishimaru, S.E. Wilson, Effect of coacervated a-elastin on proliferation of vascular smooth muscle and endothelial cells, Angi-ology 49 (1998) 289-297. 

Recombinant tropoelastin and elastin-derived peptides and proteins have also been employed to investigate the mechanism of coacervation above the transition temperamre in vitro. For the recombinant human derivative SHELA26A, the coacervation process has been demonstrated to proceed in phases that depend... 

Despite the foregoing evidence, the type-II p-tum has not been unequivocally established as the determinative element for the development of elastomeric behavior within the elastomeric domains of elastin and other native polypeptide materials. Spectroscopic analyses of native elastin and elastin-mimetic polypeptides have provided strong evidence for the presence of alternative conformations, particularly above the inverse transition temperature, which may contribute to the elastomeric behavior of the material in its coacervate state. In addition, the elastomeric domains of native tropoelastin display quite substantial sequence diversity, although it appears that Pro-Gly sequences are conserved... 

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