We have created a de novo full-length modular collagen gene that enables fabrication of specific mutants with high potential in regenerative medicine, tissue engineering and drug delivery.  Although native collagen is typically extracted from animals, safety concerns and variability limitations of animal protein drive the development of recombinant expression systems such as yeast.  In developing our modular collagen gene, we overcame the challenges of mishybridization during PCR and site-directed mutagenesis, and can mutate at defined locations, combinations and frequencies.  To face the challenge of expressing thermally stable collagen with the desired hydroxylation percentage, we developed different integrant and plasmid systems that co-express the two subunits of human prolyl hydroxylase and collagen at different gene ratios and expression levels.  We report the different hydroxylation abilities of these systems and demonstrate that the resulting full-length collagen and variant proteins can be successfully expressed, purified, and characterized.

Expression of modular collagen and variants in <i>Saccharomyces cerevisiae</i>

243rd National Meeting (2012)

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Expression of modular collagen and variants in Saccharomyces cerevisiae