The Protein Man's Blog | A Discussion of Protein Research

Protein Storage and Stability

What are Proteins?

As one of the multifarious macromolecules, proteins are complex and crucial for cellular functions. Proteins are polymers built of monomer subunits called amino acids connected by a specific type of covalent bond known as a peptide bond. The properties of the protein depend on the type of amino acids present in them. Although the primary structure of the protein comprises of the amino acid sequence, the functional properties of the protein crucially rely on the three-dimensional or tertiary structure. Similarly, protein modifications for e.g., glycosylation, phosphorylation, may change the properties and function of the protein. These modifications alter the local conformation and mediate folding or stability as well as drive proteins to different cellular compartments. Proteins also display remarkable variability in terms of structure and flexibility depending upon their folding patterns. Some proteins are relatively rigid hence can function as structural meshes or connective lines. Proteins with reversible conformational changes (polymerization or depolymerization) are crucial for protein-protein interaction, growth and the transmission of information from cell to cell or within the cell.

Affinity chomatography is a routinely used technique for purifying or enriching a protein or molecule of interest through its specific binding affinity. The target protein adheres to a particular ligand that has been immobilized on a solid support (usually beaded agarose resin). This process produces high selectivity, resolution, and capacity for the protein of interest. Traditionally, affinity chromatography is performed in column format. Where the sample is applied and eluted by gravity flow through a packed resin bed of one to several milliliters. However, gravity flow columns are time consuming and require constant attention while supernatant filters through the column and resin. This process can be accelerated with G-Biosciences’ FastPure™ Spin Columns (Mini and Midi) specifically designed for simple and efficient small scale protein purifications. FastPure™ Spin Columns combine the effectiveness of gel filtration, and the speed of centrifugation for quick and reliable protein purification.

Chromatography is a versatile field with a wide range of applications. It’s accomplished by fractionating a mixture into its molecular components. Chromatography was first used in 1901 by Russian botanist Mikhail Tsvet when he realized the technique could separate plant pigments. It has since become widely developed and utilized for separation analysis in various scientific fields.

There are several contemporary methods for protein purification. Some techniques are very powerful, such as reverse phase chromatography, immunoadsorption, or affinity chromatography. However, these methods may ultimately denature protein and negatively impact the desired product. Likewise, recombinant proteins recovered from inclusion bodies are often aggregated and nonfunctional. In all of these situations, the purified protein product must be renatured for further use.