Adjustable luciferase segmentation fusion protein and preparing method and application thereof

Abstract

The invention relates to the technical field of molecular biology, in particular to adjustable luciferase segmentation fusion protein and a preparing method and application thereof. The fusion protein comprises luciferase N-terminal protein, a flexible peptide fragment 1, MBP protein, an active valve, a flexible peptide fragment 2 and luciferase C-terminal protein, wherein luciferase N-terminal protein and luciferase C-terminal protein form luciferase together, and the active valve is a fragment of 4-8 hydrophobic short-peptide amino acids. By means of the activity adjustment mode that a short-peptide chain amino acid sequence is used as the active valve and specific cleavage enzyme is used as a valve key, a novel fluorescence complementation protein switching mode is realized, and the novel fluorescence complementation protein switching mode can be used for evaluating interaction of other protein dynamics such as combination of a ligand and a substrate, interaction capable of generating a new binding site, protein-protein interaction capable of deactivating protein, and protein-protein interaction capable of changing the specificity of a protein active substrate.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to an adjustable luciferase fragment fusion protein, its preparation method and application. Background technique [0002] With the in-depth study of subcellular structure and function, molecular physiology and pathology, and intercellular and intracellular signaling pathways, human beings' understanding of diseases and the essence of life has been traced back to the level of proteins and genes. Many life activities are realized by the combination and dissociation of protein molecules. Various important physiological activities of cells, the response of cells to the external environment and internal environment, etc., are all based on the interaction between proteins. guide the biological activity of cells and form a signal transduction network system. Therefore, an in-depth study of protein interactions is a necessary prerequisite for the recognition and understanding of...

AI Technical Summary

Problems solved by technology

However, the real-time performance of the double fluorescent complementation technology (BiFC) is poor. It requires that the two fragments can be complementary and reintegrated on the cut surface to form a complete fluorescent protein to restore the activity of the fluorescent protein. Fluorescent complementary signals can only be detected successfully in a few minutes or even hours, especially in such a complex environment in living cells, the actual situation is unknown, and protein interaction can only be assessed by changes in optical signal intensity to a great extent There may be deviations

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