Question

Explain the advantage of using protein determination in the context of modern biotechnology. Support your explanation with a current concrete example.

yes

Answer 1

ANSWER

Genetic engineering is the cornerstone of modern biotechnology. It is based on scientific tools, developed in recent decades, that enable researchers to:

ADVANTAGES

Crystallisation of proteins is a necessary step for elucidating their detailed 3-dimensional structure. Crystallisation is a difficult process that requires a highly purified protein kept in ideal conditions. In DLS measurements, polydispersity is a measure of the purity of a sample. A protein sample with a very low polydispersity indicates that it is highly purified, that all the protein is in one particular oligomeric conformation and that its structure is very well controlled under these conditions, all of which are required for crystallisation. By identifying a protein sample with the lowest polydispersity, a researcher can find the most suitable conditions for crystallisation.

In adverse conditions such as extremes of temperature and pH, a protein will become denatured. By controlling these conditions, and measuring the hydrodynamic radius, the melting point of the protein can be established. This is related to the stability of the protein and can be used as a predictor of shelf life.

MODERN PROTEIN ANALYSIS TECHNIQUES

Protein complex analysis involves extensive interpretation of the structure and function of proteins, which are present in complex biological samples. Though recent protein complex analysis methods are efficient in identifying the structure of protein complex, there are some limiting factors.

LIGHT SCATTERING

Light scattering techniques are particularly sensitive to larger molecules in preparations of smaller molecules. Any increase in the size of a protein will most likely be the result of aggregate formation. The sensitivity of the light scattering measurement to larger proteins means that the earliest stages of denaturation, leading to the formation of a few aggregates, will result in changes in the mean hydrodynamic size.

Batch Dynamic Light Scattering (DLS): Size measurement is the primary measurement of proteins that can be performed with batch-mode DLS. Since proteins have a very consistent composition and fold into tight structures, the hydrodynamic size relates predictably with molecular weight. The activity and function of a protein is closely related to correct folding and structure. As such, activity is also directly related to the size of the protein, meaning size can also be used as a predictor of activity. DLS the most sensitive technique for detecting small quantities of aggregates in preparations. Zetasizer software has a model to predict the molecular weight of a protein from its hydrodynamic size by DLS. Request a demo.

Static Light Scattering (SLS): Following on from DLS measurements, SLS measurements can also be made of proteins. Often highly purified, many protein samples should be applicable for batch measurements of molecular weight using SLS, as long as the concentrations are accurately known. By measuring the amount of light scattered at different concentrations of sample, the molecular weight, which is proportional to the amount of light scattered, can be calculated by creating a Debye plot. The slope of the line in the Debye plot is 2x the 2nd virial coefficient (a measure of molecular interaction within a solution) so this technique can also be useful for studying crystallisation conditions. A strongly positive value indicates good solubility while a strongly negative value indicates a propensity to aggregate. Request a demo.

Also use

• MULTI-DETECTION GPC/SEC
• CIRCULAR DICHROISM SPECTROMETRY
• ISOTHERMAL TITRATION CALORIMETRY