Production of recombinant human type I procollagen homotrimer in the mammary gland of transgenic mice

The large scale production of recombinant collagen for use in biomaterials requires an efficient expression system capable of processing a large (> 40 0 Kd) multisubunit protein requiring post-translational modifications. To i nvestigate whether the mammary gland of transgenic animals fulfills these r equirements, transgenic mice were generated containing the alpha S1-casein mammary gland-specific promoter operatively linked to 37 Kb of the human al pha 1(I) procollagen structural gene and 3' flanking region. The frequency of transgenic lines established was 12%. High levels of soluble triple heli cal homotrimeric [(alpha 1)(3)] type I procollagen were detected (up to 8 m g/ml) exclusively in the milk of six out of 9 lines of lactating transgenic mice. The transgene-derived human procollagen chains underwent efficient a ssembly into a triple helical structure. Although proline or lysine hydroxy lation has never been described for any milk protein, procollagen was detec ted with these post-translational modifications. The procollagen was stable in mil; minimal degradation was observed. These results show that the mamm ary gland is capable of expressing a large procollagen gene construct, effi ciently assembling the individual polypeptide chains into a stable triple h elix, and secreting the intact molecule into the milk.