Production of Synthetic Methionine-Free and Synthetic Methionine-Limited Alpha Casein: Protein Foodstuff for Patients with Homocystinuria due to Cystathionine Beta-Synthase Deficiency

The aim of this study was to evaluate the possibility that synthetic forms of methionine-free alpha-casein and methionine-limited alpha casein could be produced by recombinant means to form the basis for developing an industrial-scale process for the provision of a foodstuff suitable for patients with homocystinuria due to cystathionine beta-synthase (CBS) deficiency. As a first step, two forms of alpha casein gene, encoding methionine-free alpha casein (Fcas) or a methionine-limited alpha casein (Mcas), were synthesised and expressed in Escherichia coli. Using the overexpression vector pET28a, both genes were highly expressed in E. coli in soluble form as well as in inclusion bodies. The two recombinant proteins were purified by the one step methods using the fused His-tag and the Ni²⁺column and validated by Western blot analysis. This work paves the way for industrial-scale production of proteins suitable for patients with homocystinuria due to CBS deficiency.