Protein purification using anion exchange membranes - Term Paper Example

This change results in the protein being less capable of forming a strong ionic interaction with negatively charged cation exchanger. A similar chain of events occurs with anion exchange media. At a lower pH of the mobile phase the target molecule becomes more protonated and hence positively charged. The result is that the target molecule no longer has the capability to form a strong ionic interaction with the positively charged anion exchanger which causes the molecule to elute from stationary phase.

Ion exchange is the most commonly practiced chromatographic method of protein purification due to its ease. This technique exploits the amphoteric character of a protein ( net positive in low pH buffer and negative in a high pH buffer). The technique exploits the fact that the distribution and net charge on the protein’s surface determines the interaction of the protein with the charged groups on the surface of the immobile phase, an anion exchange membrane in this case. The charges on the protein and the membrane must be opposite for the exchange interaction to occur.

The support membrane, which has covalently attached positive functional groups, is referred to as an anion exchanger if mobile negatively charged anions will be the exchanged species. . In membrane chromatographic processes, the transport of solutes to their binding sites take place predominantly by convection and the pore diffusion is very small comparing with the beads column, thereby the mass transfer resistance is tremendously reduced. Membrane chromatography is a promising process for the isolation, purification, and recovery of proteins, enzymes, and nuclear acids.

Comparing with traditional beads column chromatography, membrane chromatography can be faster, easier and cheaper to mass-produce. And also, it is easy to set up and scale up. Most important is saving time in membrane chromatography, this is important because many proteins lose their activities with time. Membranes are also more convenient because they do not require column hardware or packing, they reduce buffer usage and floor space requirements and they generally improve manufacturing flexibility.

There are mainly three shapes of membrane available, including flat sheet, hollow fiber and radial flow used for protein purification (2). (From http://www.natrixseparations.com/media/application_note7.pdf) Figure 1 Principles of an anion exchange chromatography: Lowering pH on the protein elution from positively charged anion-exchange membrane. Molecules with higher charge density bind much stronger to the membrane and consequently require greater change in pH to be released from the membrane surface.

Theory: Proteins are bound to anion exchange membranes by reversible, electrostatic interactions. A separation is obtained because the diverse array of molecular species have different affinities for the exchanger. The adsorbed proteins are eluted in order of least to most strongly bound molecules,