Petroleum Jelly – What It Is And How It Is Made

Proteins can be modified and designed to fulfil the needs of special applications related to the biotech industry. These protein engineering techniques are widely used in biotechnology research in an effort to re-design or modify proteins.

Scientists use these techniques to isolate proteins before they undergo a purification process. Purification of isolated proteins is done in order to facilitate proper studies on the specifications and conformations of the substrate. It also allows reactions between proteins and other ligands to be studied as well as enzyme-specific activities. Scientists refer to ligands as proteins that become attached to special receptor proteins.

The degree of purity during the purification process will depend mainly on the demands for a particular protein. In some instances, for example, a simple crude extract will be sufficient. A higher level of purification may be needed for applications such as the food industry or for pharmaceutical purposes. Several purification techniques are therefore required in order to attain the purity level desired.

Developing a Strategy

During each step of the purification process, the content of the product is reduced. Ideal purification strategies, therefore, involve those that will produce the highest level of purification in the least number of steps. However, the steps used during the purification process are dependent upon the size, charge, solubility, and other properties of the particular protein being used. The following procedures are therefore more suited for the purification of single-cytosolic proteins. (Different techniques are required in the case of cytosolic protein complexes).

• Preparing a Crude Extract

Purifying intracellular proteins that exist inside a cell is the first major step in preparing a crude extract, and can be done using a double diaphragm pump. Crude extracts commonly contain a highly complex blend of proteins derived from the cytoplasm in the cell along with nutrients, macromolecules, and co-factors.

Once the crude extract has been prepared it can be used for several different applications in biotechnology. If purity becomes problematic, a subsequent set of steps must be undertaken to achieve the level of purity desired. In order to prepare the protein extracts a process of cellular debris removal is undertaken. These extracts are created by cell lysis, accomplished by using chemicals, enzymes, sanitation, or a French Press.

Removing Debris from the Crude Extract

A process of centrifugation is used to remove debris from the crude extract. After completion of the centrifugation process, the supernatant can be successfully recovered. With extracellular preparations, proteins can be naturally obtained during the centrifugation process when cells are removed.

In some biotechnology practices, thermostable enzymes are required. These are enzymes that are capable of withstanding the highest temperatures without becoming denatured. They are also capable of maintaining a high level of activity.

Intermediate Protein Purification Steps

Most modern biotechnical protocols have so far made use of commercially obtainable kits. In addition, for standard procedures, these protocols often provide techniques that offer a more ready-made solution. When conducting protein purification activities, filters and gel-filtration columns are commonly used.

Dialysis Kits

When using a dialysis kit it is important to follow the exact directions provided by the manufacturer of the kit. Following the instructions will ensure that the right amount of solution is used and the appropriate waiting time is observed. The waiting time specified is there to ensure that the right concentration of fluid is reached before it enters a clean test tube.

Chromatographic Methods

The chromatographic method makes use of either bench-top columns or automated HPLC equipment. With HPLC equipment, ion-exchange, size-exclusion, or reverse-phase methods are used. Diode array or laser technology is used to detect samples.