The interest to produce gelatins by recombinant methods is ever increasing. The widespread use and possibilities for use of gelatins in medical and clinical applications poses higher demands on gelatins in every aspect, starting by the need for providing processes that are economically viable to produce gelatins. In turn, this need has prompted careful consideration of process variables and variations in protein sequences that could influence expression properties, and hence yields of desired gelatins that could be obtained.
EP 926543 and Werten et al. 1999 (Yeast 15, 1087-1096) describe a production method of recombinant gelatins, wherein high yields of non-hydroxylated fragments of the helical domain (consisting of Gly-Xaa-Yaa triplet repeats) of mouse type 1 (encoding a 21 kDa and 28 kDa, calculated MW, COL1A1 peptide and a 53 kDa COL1A2) and rat type III (COL3A1) are produced in the methylotrophic yeast Pichia pastoris. A factor such as the fermentation pH was in certain cases found to be of influence on stability of the expressed product, but also the presence proteolytic sequences in the expressed gelatin was found to be relevant and it has been hypothesized that similarity of codon usage with endogenous proteins of the host microorganism could be relevant for obtaining high yields of desired exogenous proteins.
In US 2006/0241032 XRGD-enriched gelatin-like proteins with a minimum (increased) level of XRGD motifs and with a certain distribution of said XRGD motifs are disclosed that were found to be highly suitable for cell adhesion and cell binding in medical and biotechnological applications. The cell binding peptides described therein have good cell attachment properties.
In order to be susceptible for applications of the recombinantly produced gelatins in human beings a lot of effort was directed to make recombinant gelatins as close as possible resembling human gelatin.
However, susceptibility to degradation has been a limiting factor in the ability to produce large amounts of recombinant gelatins.
USPTO Backgrounds
USPTO Backgrounds