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Yeast. 2001 Jun 30;18(9):797-806.

High-level production of human type I collagen in the yeast Pichia pastoris.

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Collagen Research Unit, Biocenter and Department of Medical Biochemistry, PO Box 5000, FIN-90014 University of Oulu, Finland.


Four human genes, two of them encoding the proalpha1 and proalpha2 chains of type I procollagen and two of them the two types of subunit of prolyl 4-hydroxylase (4-PH), were integrated into the genome of Pichia pastoris. The proalpha1 and proalpha2 chains expressed formed type I procollagen molecules with the correct 2:1 chain ratio, and the 4-PH subunits formed an active enzyme tetramer that fully hydroxylated the proalpha chains. Chains lacking their N but not C propeptides formed pCcollagen molecules with the 2:1 chain ratio and, surprisingly, the expression levels of pCcollagen were 1.5-3-fold relative to those of procollagen. Both types of molecule could be converted by pepsin treatment to collagen molecules that formed native-type fibrils in vitro. The expression levels obtained for the pCcollagen using only single copies of each of the four genes and a 2 l fermenter ranged up to 0.5 g/l, indicating that it should be possible to optimize this system for high-level production of recombinant human type I collagen for numerous medical applications.

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