October 27-30, 2001
X National Congress of Biochemistry and Molecular Biology of Plants. La Paz, Baja California Sur, Mexico.

Use of Transgenic Plants for the Production of Structural Proteins. Julio Baez, FibroGen.

Several complex eukaryotic structural proteins (silk, elastin, collagen) are under development for pharmaceutical applications as biocompatible materials. For most of these applications, over 100kg/year could be required to be produced at less that $US100/g. For these reasons transgenic systems are under development for these proteins. This presentation will describe the use of transgenic plants for the production of one of these mammalian proteins: gelatin. It is currently produced from denatured and hydrolyzed animal carcass derived collagen. Gelatin is widely used in the pharmaceutical industry for capsules, as vaccine stabilizers, and as plasma expanders. There are concerns related with the safety and quality consistency of animal-derived gelatin due to the potential for contamination by prions, product heterogeneity, and the adverse immunological reaction.

We will discuss the expression of human collagen III in tobacco cells used as a model for establishing the feasibility of producing hydroxylated collagen in plants for gelatin production. Tobacco cells were engineered to accumulate full length pro human Collagen (III) and at the same time express the human alpha and beta subunits of prolyl 4 hydroxylase. Expression was regulated by a constitutive promoter. Nuclear transformation via a gene gun was carried out, and the clones expressing the three proteins were selected by northern and western blotting. Selected clones were cultivated in shake flasks to generate cell mass from which pro collagen III was purified to a homogenous state for analysis of hydroxylation of proline. In tobacco cells, all three genes were transcribed and translated into protein with the alpha and beta subunits forming an active prolyl hydroxylase capable to hydroxylate prolyl residuals on collagen chains. This technology could be used to produce genetically designed plant derived gelatin cost-effectively to meet the specific requirements of various gelatin applications and to also address shortages of certain types of gelatin.