1. Determination of primary structure of protein by primary structure. Also known as protein sequence analysis, the basic methods are as follows: 1. The chemical chain cleavage method and protease hydrolysis method are used to specifically cleave the polypeptide chain. 2 The order of each purified small peptide was determined one by one. 3 Determine the order of arrangement of small peptide segments based on the overlapping regions in the amino acid sequence of the peptide. 4 Complete the sequence analysis of the entire polypeptide chain. Although protein sequence analysis has been automated, it is still time consuming, complicated, and expensive. After the emergence of recombinant DNA technology, one can directly derive the amino acid sequence of a protein from a cDNA or gene sequence, which is fast and economical, and has become the most commonly used method for determining the primary structure of a protein. 2, protein high-order structure prediction protein three-dimensional structure largely determines the function of the protein, the application of X-ray crystal diffraction pattern and nuclear magnetic resonance method has determined the structure of more than 10,000 kinds of proteins and their complexes, but with the measured Compared with more than 300,000 protein sequences, there is still a big gap, which greatly affects the study of the relationship between protein structure and function. Therefore, it is particularly important to develop a protein structure prediction method that is independent of experiment and has certain accuracy. In recent years, many important achievements have been made in this respect. The theoretical prediction methods of protein structure are based on the theory that the primary structure of amino acids determines the high-level structure, and are roughly divided into the following three categories. (1) Comparative modeling method, also known as homology structure prediction, is a protein structure whose structure is known to be unknown and whose structure is unknown based on a large number of known three-dimensional structures of proteins. (2) Reverse folding method The reverse folding method is a relatively new method developed in recent years. It can be applied to the case where there is no homologous structure, and there is no need to predict the secondary structure, and the tertiary structure can be directly predicted, so that the prediction accuracy of the secondary structure at the current stage is not more than 65%, which is a potential. Forecast method. (3) De novo prediction method In theory, the de novo prediction method is the most ideal protein structure prediction method. It requires that the method itself can predict the secondary structure and higher structure of the protein based only on the amino acid sequence of the protein, but it is not yet fully met. According to the data in the database established by the analysis of the structure and function of natural proteins, the spatial structure and biological function of peptide chains of certain amino acid sequences can be predicted; on the contrary, the amino acid sequence and spatial structure of proteins can be designed according to specific biological functions. Through the experiments such as genetic recombination, the relationship between structure and function can be directly examined. 3, protein three-dimensional structure experimental determination According to the state of the protein, the method for determining the three-dimensional structure of proteins is divided into two categories: 1 using X-ray crystal diffraction pattern method and neutron diffraction method to determine the protein molecular conformation in the crystal. 2 Determination of protein conformation in solution by nuclear magnetic resonance, circular (circular) dichroism spectroscopy, laser Raman spectroscopy, fluorescence spectroscopy, ultraviolet differential spectroscopy and hydrogen isotope exchange. Other methods can determine the local conformation of protein molecules in solution, but it is difficult to obtain a complete three-dimensional structure of protein molecules, which has great limitations in application.
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