Methods of producing proteins having triple-helix structure

摘要

An objective of the present invention is to provide methods of producing human collagen molecules that are easy to isolate and purify and that have a structure substantially equivalent to that of a natural collagen molecule, wherein host cells that are transduced with a collagen gene synthesize large amounts of human collagen protein derived from a gene introduced into a high exogenous gene expression vector. Another objective of the present invention is to provide collagen molecules produced by the production methods.;The present inventors discovered that a large amount of human collagen hardly contaminated with host cell-derived collagen could be produced, by selecting from various mammalian cells a host cell that has low collagen expression and introducing a collagen gene construct into a vector capable of high exogenous gene expression.

主权项

1. A method of producing a protein having a triple-helix structure, wherein the method comprises:
(a) introducing DNA encoding a protein having a triple-helix structure into a pNOW/CMV-AA vector; (b) transforming an isolated cell with the vector, wherein the isolated cell is a Chinese hamster ovary (CHO) cell to thereby provide a transformed cell; and (c) culturing or breeding the transformed cell, and (d) collecting a protein having a triple helix structure from the transformed cell or culture supernatant thereof, thereby obtaining an expression level of 50 μg/mL or more of the protein having a triple helix structure is selected from the group consisting of human type I collagen, human type II collagen, and human type III collagen or a partial peptide thereof.