Proteome analysis - what is required for identification!
Proteins are the all-controlling ‘building blocks of life’. The basic structure, the polypeptide chain, is based on the genetically determined sequence of 20 different amino acids, which occur in different frequencies:
In the next step, this basic structure is changed by so-called post-translational modifications (PTM), by splitting off some amino acids and various chemical changes that are responsible for the final function of the protein.
While the basic structure of the amino acid sequence can be deduced from the genome, this is not possible for the post-translational modifications (PTM). Knowledge of the PTM is essential for determining the function of a protein. Depending on the PTM, proteins from the same backbone can accelerate, slow down, completely block or not influence certain processes at all. They ‘embody’, to put it bluntly: ‘permanent dynamics and change’ - just like life!
The biological function in its complexity of a single protein consisting of 6 forms:
Biological functions of an example protein, Pac1, of which differently modified variants with different biological functions occur. The hypothetical protein Pac1 has little biological activity as a polypeptide chain and is only converted into its active form, Pac1A1, by phosphorylation and glycosylation. The protein can also be specifically cleaved by proteases into Pac1A, which fulfils an alternative function, Pac2A and B, which inhibit the function of active Pac1A1.
In order to determine the role of Pac1 in the respective diseases, all forms of Pac1 must be analysed in detail. Mixing several forms of the protein inevitably leads to incorrect results that cannot be interpreted.
Mosaiques analyses the proteins and peptides actually present in their unchanged form. The analysis of mosaiques is direct, i.e. the concentration of the protein is measured directly in the mass spectrometer. With this technique, all 6 forms of Pac1 can be detected independently.
Alternative mass spectrometry-based technologies measure fragments (typically tryptic peptides) that are previously generated from the proteins in the sample. The amount of specific peptide is then used to infer the original protein present, with different results depending on which fragment is analysed. Modifications to proteins, which are crucial for their function, are not taken into account.
In the present example of Pac1, the other mass spectrometric techniques, which do not correspond to mosaiques' unique technique, would provide information on the sum/combination of Pac1, Pac1A and Pac2A, but would not detect the actual active form Pac1A1.
Non-mass spectrometry-based techniques utilise ‘probes’, e.g. antibodies or aptamers, which bind a certain protein more or less specifically. The quantity of the bound molecules is used to deduce the quantity of the specific protein. This also generally does not provide any information about post-translational modifications to the protein. The specificity is often not defined and it is often unclear whether the bound protein is actually the target protein or another, similar protein.
In the present example of Pac1, this technique would provide information about the sum/combination of Pac1, Pac2A and Pac2B, but would also not detect the actual active form Pac1A1.
To determine the amount of active Pac1A1 present and at the same time the amount of Pac1 variants with different functions, only the Mosaiques technology is suitable and has been proven. This information can be found across all samples from the clinical studies and all measured proteins in the database, including their changes over the course of the disease. Only the high accuracy of all the information in the proteome makes it possible to compare and identify the proteins involved in the disease process to be determined, such as cardiovascular, kidney or a specific cancer. This has made it possible for the first time to recognise and define the disease-specific proteome as a whole. This usually contains several hundred proteins, while the individual patient usually only forms parts of it. If this patient has not formed the target proteins of the administered drug in his disease proteome, the drug cannot work. On the one hand, proteome analysis makes it possible to recognise the chronic disease at an early stage before organ damage occurs and, on the other hand, it can precisely adjust the patient with the drug(s) acting on his proteome. This is one of the reasons why the drugs have so far only been effective in 15-30% of patients. This is also due to the late detection of the disease.
The extensive medical histories are also contained in the database on the course of the disease with their specific additional proteome biomarker patterns. The data comes from over 100 extensive clinical studies involving over 1,200 renowned doctors and scientists. These scientists and physicians are autonomous, independent and have raised the funding for the clinical proof-of-concept studies themselves. This is unique in the world and demonstrates the unrivalled seriousness and credibility of proteome pattern analysis by mosaiques and its subsidiaries, protexam and xken-health.
Every proteome analysis is carried out in the same way, whether it is just to determine several diseases or to check the effect of medication. This makes the individual precise findings more expensive than a test for a single molecule, which can only be obtained for a few euros and often provides false results (see PSA or albuminuria). In most cases, it is the cause of many unnecessary invasive further investigations, such as biopsies or even unnecessary surgery. Modern medicine requires precision in favour of the patient's health!