Design and bioproduction of a recombinant multi(bio)functional elastin-like protein polymer containing cell adhesion sequences for tissue engineering purposes

Abstract

Genetic engineering techniques were used to design and biosynthesise an extracellular matrix (ECM) analogue. This was designed with a well-defined molecular architecture comprising different functional domains. The structural base is a elastin-derived repeating unit, which confers an adequate elastic characteristic. Some of these elastin domains have been modified to contain lysine; this amino acid can be used for crosslinking purposes. The polymer also contain periodically spaced fibronectin CS5 domains enclosing the well-known cell attachment sequence REDV. Finally, the polymer has target sequences for proteolitic action. These sequences are those found in the natural elastin and are introduced to help in the bioabsorption of the polymer. In addition, these proteolitic sequences were chosen in a way that, after proteolitic action, the released fragments will be bioactive. These fragments are expected to promote cell proliferation activity, angiogenesis and other bioactivities of interest for tissue growing, repairing and healing. After purification, the resulting polymers proved to be of high purity and correct sequence. Glutaraldehyde has shown to be a cross-linking agent for this polymer, yielding insoluble hydrogel matrices. This work is framed in a long term project aimed to exploit the power of genetic engineering for the design and bioproduction of complex ECM analogues showing the rich complexity and multi (bio)functionality of the natural matrix.