Abstract
The purpose of the present study was to evaluate de influence of protein–sugar complexation on the stability and functionality of C-reactive protein, after exposure to constant high temperatures, in order to develop highly stable positive controls for in-vitro diagnostic tests. C-reactive protein is a plasmatic protein used as a biomarker for the diagnosis of a series of health problems such as ulcerative colitis, cardiovascular diseases, metabolic syndrome, due to its essential role in the evolution of chronic inflammation. The sugar–protein interaction was investigated using steady state and time resolved fluorescence. The results revealed that there are more than two classes of tryptophan, with different degree of accessibility for the quencher molecule. Our study also revealed that sugar–protein complexes have superior thermostability, especially after gamma irradiation at 2 kGy, the protein being stable and functional even after 22 days exposure to 40 °C.
Highlights
The purpose of the present study was to evaluate de influence of protein–sugar complexation on the stability and functionality of C-reactive protein, after exposure to constant high temperatures, in order to develop highly stable positive controls for in-vitro diagnostic tests
A concentration lower than 3 mg/L is normal for a healthy adult, values between 3 and 10 mg/L mean a minor increase that can be associated with obesity, pregnancy, depression, diabetes, common cold, gingivitis, sedentary lifestyle, smoking or genetic polymorphisms, while values between 10 and 100 mg/L are associated with systemic inflammation such as rheumatoid arthritis and lupus erythematosus, or other autoimmune diseases, malignancies, myocardial infarction, pancreatitis or bronchitis
We tested, using the obtained data, the method of analysis of fluorescence quenching using the modified Stern–Volmer plots[15], plotting the variation of (F0/F0 – F) dependence by (1/[Q]), and within this analysis method the obtained graph showed a downward curvature. This implies that there are more than two classes of tryptophan molecules with different degrees of accessibility for sucrose, C-reactive protein (CRP) protein having in his primary structure six Tryptophan r esidues[17]
Summary
The purpose of the present study was to evaluate de influence of protein–sugar complexation on the stability and functionality of C-reactive protein, after exposure to constant high temperatures, in order to develop highly stable positive controls for in-vitro diagnostic tests. This protein plays an essential role in the evolution of chronic inflammation, and is the result of constant deterioration of the inner walls of the arteries. A series of rapid tests are been marketed for the detection/quantification of CRP, used as a biomarker for the assessment of various disorders or the risk of developing them They can offer information about the active phase of autoimmune conditions such as Crohn’s or ulcerative colitis, the risk of developing a cardiovascular disease and more recently studies had shown that this protein is very important in the diagnosis of the Metabolic Syndrome (MS). The present study aims to establish how the addition of sucrose in a CRP solution can improve the thermostability of the protein and the optimal concentration for its active maintenance, even after prolonged exposure to 40 °C ± 1 °C
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