Process for purification of a fatty acid binding protein
A process for purification of a fatty acid binding proteins such as, e.g., Sm14 of Pichia pastoris or type-3 FABP protein of Fasciola hepatica . The process includes the steps of: (a) performing lysis of cells containing the fatty acid binding protein to obtain a lysate; (b) clarifying the lysate obtained in step (a) to obtain a clarified lysate; (c) loading the clarified lysate in a column containing an anion exchange resin; (d) eluting proteins from the column by pH changes in the column; and (e) separating contaminant proteins from the fatty acid binding protein by gel-filtration.
1. A process for purification of a recombinant Sm14 Schistosoma mansoni protein, said process comprising the steps of:
(a) performing lysis of cells containing the Sm14 protein to obtain a lysate;
(b) clarifying the lysate obtained in step (a) to obtain a clarified lysate;
(c) loading the clarified lysate in a column containing an anion exchange resin;
(d) eluting proteins from the column by pH changes in the column; and
(e) separating contaminant proteins from the Sm14 protein by gel-filtration, wherein said Sm14 protein is encoded by SEQ ID NO:3 and is expressed in Pichia pastoris.
2. The process according to claim 1 , wherein the lysis performing step comprises resuspending the cells in 30 mM Tris-HCl 30 mM pH 9.5 buffer and lysing the cells with a French press.
3. The process according to claim 2 , wherein the clarifying step comprises centrifugation.
4. The process according to claim 3 , wherein the loading step comprises loading the column with the clarified lysate in 30 mM Tris-HCl pH 9.5 buffer.
5. The process according to claim 4 , wherein the eluting step comprises eluting the fatty acid binding protein from the column with 30 mM Tris-HCl pH 8.0 buffer.
6. The process according to claim 5 , wherein the anion exchange resin comprises a quaternary amine in a matrix of cross-linked agarose with dextran surface extenders.
7. The process according to claim 6 , wherein the separating step comprises gathering and concentrating eluted fractions from the eluting step, applying the eluted fractions to a gel filtration column containing size exclusion media comprised of the anion exchange resin allyl dextran and N,N′-methylene bisacrylamide, using phosphate buffered saline pH 7.4 as a mobile phase.