Polymer substrate for recombinant protein purification and environmental remediation

Medicilon's protein scientists have been working on protein expression and purification for many years. We can start your project even you have nothing in hand but the name of your protein. In Medicilon's laboratories, protein purification is performed in scales from micrograms and milligrams. All Protein Purification Services start with the analysis of physico-chemical and biological properties of a target protein resulting in the development of tailored procedures for its extraction, purification and characterization. Email:marketing@medicilon.com.cn Web:www.medicilon.com

A polymer substrate for recombinant protein purification is provided. A process for purifying recombinant proteins is also provided. In general, the present invention is directed to a chelating composition which may be used in a self-supported purification medium for recombinant protein purification. In another embodiment, a process for removal of transition metals from environmental waste is also provided.

While recombinant protein development is vital to areas in the biochemical field, an efficient and effective method for recombinant protein purification remains a major obstacle. Existing methods typically involve metal ion affinity chromatography; these methods, however, have many limitations.

Such existing methods of recombinant protein purification typically involve some form of column-based separation and, as such, require resins that are both expensive to manufacture and to purchase, and are only suitable for small-scale laboratory operations due to the inherent nature of column based separations, giving rise to significant protein loss and the recovery of only small quantities. Additionally, current methods for larger scale purifications must be achieved by a series of buffered extractions and centrifuging. Such large scale processes can be solvent intensive and do not necessarily give rise to pure products or significant yields. Methods for large-scale, efficient purification of selected proteins are essential to the pharmaceutical industries as well as other biochemistry-based concerns.

A need thus exists for an efficient method for purification of recombinant proteins. Ideally, such a purification medium would be self-supported, inexpensive to manufacture, and as effective as existing methods. In addition, it would be desirable to have a purification medium that is easily recovered and could be configured for use in continuous processes in any desired structure. Finally, it would be desirable if such a medium could also be used in environmental remediation of heavy metals.

Various features and advantages of the invention will be set forth in part in the following description, or may be obvious from the description, or may be learned through practice of the invention.

Generally, in one embodiment, the present invention is directed to a self-supported purification medium for recombinant protein purification and to a process for using that medium. A self-supported purification medium does not require a column or special machinery and, therefore, is inexpensive to manufacture while retaining the effectiveness of existing methods. In addition, a self-supported purification medium is easily recovered and can be configured for use in continuous processes in any desired structure. The purification medium may comprise a polymer substrate, a chelating agent that is formed from the reaction of the substrate with a dianhydride precursor, and a transition metal ion having a 2+ oxidation state. The polymer substrate may contain amine groups or hydroxy groups. The amine groups or hydroxy groups of the polymer substrate are reacted with the dianhydride precursor to form the chelating groups that are covalently bonded to the substrate. The chelating substrate is then complexed with the transition metal ion having a 2+ oxidation state. The self-supported purification medium is next contacted with a composition containing a histidine-tagged protein. The histidine-tagged protein binds with the chelated transition metal ion having a 2+ oxidation state thereby separating the histidine-tagged protein from the composition.

In certain embodiments, the chelating agent precursor may comprise EDTA, dianhydride (ethylenediaminetetraacetic dianhydride). In other embodiments, the chelating agent precursor may comprise DTPA, dianhydride (diethylenetriaminepentaacetic dianhydride). In certain embodiments, the polymer substrate may be chosen from a group consisting of PVOH (polyvinyl alcohol), cellulose, a derivative of cellulose, a polyester, Tencel® (Tencel®) is the brand name of Tencel® Ltd. for lyocell, the fiber's generic name), nylon, wool (keratin) or cotton. Also, in certain embodiments, the polymer substrate may be a fiber while in other embodiments the polymer substrate may be a film. In certain embodiments the fiber may have a diameter of less than 100 microns and may be considered a standard fiber or micro fiber while in other embodiments the fiber may have a diameter of less than 1 micron and will be considered a nano fiber. In certain embodiments the polymer substrate may be a non-woven, woven, knitted fabric, film or a polymer nanoparticle. The polymer substrate may be porous, fibrillated, etched or treated in such a way as to give rise to an enhanced surface area.

In preferential embodiments of the present invention, the transition metal ion may be chosen from Co2+ or Ni2+. In other embodiments, the transition metal ion may be chosen from Fe2+, Cu2+, and Zn2+. Still in further embodiments, any transition metal with affinity for the chelating substrate may be used.