Protein purification is a complex and non-standardized process; the fact that proteins have different structural types making it difficult to create a standard methodology to obtain them in a pure, soluble, and homogeneous form. The present study shows the selective development of a buffer suitable for proteins of interest that allows high concentrations of hGPN2 protein to be obtained with low polydispersion and high homogeneity and purity. By taking the different reagents used in the construction of different buffers as a basis and performing purifications using different additives in different concentrations to determine the optimal amounts, the developed process helps to minimize the bonds, maintain solubility, release the proteins present in inclusion bodies, and provide an adequate environment for obtaining high concentrations of pure protein. GPN proteins are of unknown function, have not been purified in high concentrations, and have been found as part of the RNA polymerase assembly; if they are not expressed, the cell dies, and overexpression of certain GPN proteins has been linked to decreased survival in patients with invasive ductal carcinoma breast cancer types ER+ and HER2+. The results of the present study show that the use of the buffer developed for recombinant hGPN2 protein expressed in Escherichia coli could be manipulated in order to isolate the protein in a totally pure form and without the use of protease inhibitor tablets. The resulting homogeneity and low polydispersion was corroborated by studies carried out using dynamic dispersion analysis. Thanks to these properties, it can be used for crystallography or structural genomics studies.
La enzima lactato deshidrogenasa o LDH, es el producto final del metabolismo presente en todas las células del cuerpo, esta puede ser detectada una vez que pasa al espacio extracelular a causa de un estado patológico como lesiones tisulares o necrosis. Se produce en distintos órganos del cuerpo, bajo cinco formas isoenzimáticas diferentes LDH1, LDH2, LDH3, LDH4 y LDH5. Se realizó una revisión bibliográfica con el objetivo de establecer la validez de la lactato deshidrogenasa como un predictor de mortalidad mediante la correcta medición de esta enzima en el laboratorio, y a su vez brindar información al personal de salud sobre su adecuada interpretación. El método estandarizado permite medir esta enzima, a través, del método fundamental de seguimiento continuo, medimos la desaparición de NADH, para ello se requiere de 2 reactivos, el piruvato y NADH, adicionalmente se requiere que el pH sea de 7,4. La longitud de onda que se debe utilizar para la lactato deshidrogenasa es de 340nm. A partir del año 2020 se empezó a utilizar la LDH como un indicador del desarrollo de una enfermedad grave y como un predictor de mortalidad en pacientes con covid-19, gracias a que se comprobó la existencia de una relación entre los niveles elevados de LDH y un peor pronóstico en pacientes con esta patología. Aunque existen factores adversos que pueden modificar los niveles de LDH, esto no imposibilita utilizar la enzima como predictor de mortalidad.
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