Importance of Protein Analysis & the Assay Methods
Protein is most vital biochemical molecule which form the body mass of living beings mostly animals.
These proteins exist in dissociated forms like amino-acids, peptide, polypeptide, enzyme, hormone or protein molecules as such as per the body and physiology requirements.
They carry out various functions in the body and almost all the body reaction occur with the involvement of protein.
They exist as single chain molecule, as 3 dimensional structures or even in bundle or complex forms.
Protein analysis & quantification:
Protein is indicator of biological entity or activity. And hence in any biological reaction or estimation or bio-process, protein analysis and quantification is done to determine the quantity, quality of protein and there by the state of biological reaction or process.
→ For instance, protein analysis or estimation is done to know the concentration of protein content in nutritious foods (Ex; protein powders, meat etc),
→In biochemical estimations of bio-samples to know their quantity (Ex; hemoglobin analysis in blood in case of anemia best range is 12 to 14 mg and in anemia the level goes down),
→In estimation of microbial contamination, to estimate the growth of microbes in culture or fermentation broths, in genetic engineering etc.
→Protein analysis and quantification is also done in times of crime or the forensic examination etc.
Methods of Protein analysis:
Protein analysis is of different methods like
Protein sequence analysis
Protein binding analysis
Crude protein analysis etc. this also means quantification of protein
All the above methods are intended for different purposes of research
For example protein sequence analysis is necessary to analyze the sequence of amino acids or peptides in a proteins like Insulin. Insulin was previously used from animals like pigs, cattle for diabetes treatment. But these insulin’s had side-effects due to difference in the amino acids in the protein sequence. Hence biotechnology came up to solve the problem by providing an Insulin which has exact sequence to human insulin with no side effects.
Protein structure analysis, proteins have different structures like secondary, tertiary, Quaternary, 3 fold structure etc. These structures influence the function of protein for example 3 dimensional ones can carry some substances by either binding them over the surface or by encasing inside their structure etc. Chain and simple structure protein easily pass through cell walls .
This can be done by NMR and X-Ray crystallography.
Crude protein analysis
This is done in terms of qualitative terms i.e to know nature and type of protein. Then it is also done in quantitative terms whereby the amount of protein in given biological sample is known.
Protein analysis in qualitative terms is done by biuret test, ninhydrin test etc.
But for quantification of protein, Lowry method is widely followed. This is a colorometric procedure where in protein sample is treated to produce a dense color as a function of protein and is then estimated by using spectrophotometry for determination of intensity which is then calculated to quantity of protein.
Initially biochemicals like enzymes, hormones etc were expressed in terms of quantity per gram of tissue or biological sample.
But this has the disadvantage that water content forms 80% of bio matter and the chances of variation are much. Hence the expression in terms of quantity per gram of protein was adopted and has least variance.