Modification of Recombinant Human Collagen with Matrix Metalloproteinase Cleavage Sites
- Author(s): Crakes, Dale;
- Advisor(s): Wang, Szu-Wen;
- Da Silva, Nancy A
- et al.
Collagen is the most abundant component of the extracellular matrix, and recombinant collagen is commonly used in tissue engineering applications. Matrix metalloproteinases (MMPs) are a specific family of enzymes that can cleave and degrade collagen. Control of the degradability of a tissue engineered matrix is an important attribute, and the modulation of MMP-cleavable sites in collagen could lead to better control over the digestibility and ease of cell infiltration in a collagen matrix. Using a computer-optimized, modular collagen III gene, recombinant collagens were designed in which native MMP-cleavable sites were deleted, added, and moved along the length of the collagen molecule, and then expressed in Saccharomyces cerevisiae. MMP-1 was used to digest each of the collagen variants, and more complete digestion was seen in collagen variants featuring more MMP sites. Hydrogel formation featuring recombinant collagen was explored, but future effort is still needed before consistent hydrogel formation can be obtained. EDC / NHS chemistry was used to covalently stabilize our recombinant collagen, which has slightly decreased thermostability compared to human collagen. Cell adhesion assays were used to confirm the increased stability of the collagen and allow cell-based experiments to be performed at 37 °C.