Composite stabilizing agent and enzyme activity determination method of NMN transferase with composite stabilizing agent

Abstract

The invention provides a composite stabilizing agent and an enzyme activity determination method of an NMN transferase with the composite stabilizing agent. The method comprises 1, preparing a crude enzyme liquid, 2, carrying out split charging on the crude enzyme liquid through multiple centrifuge tubes, adding a protective agent into one part of the centrifuge tubes and adding double distilled water into the other part of the centrifuge tubes, 3, adding a buffer solution, a substrate and metal ions into the two parts of the centrifuge tubes, uniformly mixing the materials, putting the centrifuge tubes with the mixtures into a water bath kettle, carrying out a reaction process, adding EDTA into the reaction systems, carrying out a reaction process, carrying out centrifugation, removing precipitates and collecting the supernatant, 4, taking the supernatant and determining a light absorption value of the reaction product and 5, determining relative enzyme activity through a control group which is a crude enzyme liquid without the protective agent. The composite stabilizing agent comprises common simple ingredients, has a low cost, can effectively improve NMN transferase stability and has a wide application prospect. The enzyme activity determination method of the NMN transferase is simple and practical and has visual effects.

Description

technical field [0001] The invention relates to the field of microbial engineering, in particular to a composite stabilizer and an enzyme activity assay method for NMN transferase added with the composite stabilizer. Background technique [0002] NMN transferase (Nmnat) plays a very important role in the life activities of organisms. It is a kind of protein that contributes to the complex assembly of various proteins, protein folding and protein refolding after protein damage. It can be combined with adenosine triphosphate during protein folding. (ATP) coupled, using nicotinamide mononucleotide (NMN) to generate nicotinamide adenine dinucleotide (NAD) and NAAD, maintaining the balance of NAD in the body; it is also an indicator protein and a stress response protein that can As the target of biopharmaceuticals and regulate the transcription and synthesis of target genes. A large number of studies have shown that NMN transferase has a wide range of sources in nature, mainly p...

AI Technical Summary

Problems solved by technology

However, chemical modification mainly covalently modifies certain residues on the enzyme protein peptide chain, which will cause changes in enzyme activity; although the stability of the enzyme is increased after immobilization treatment, it is easy to separate from the reaction and can be used repeatedly. It is convenient for transportation and storage, and is conducive to automatic production, but the activity of the enzyme is reduced, and the scope of use is reduced

Therefore, the most commonly used method for stabilizing enzymes is to add a protective agent, but adding a protective agent to NMN transferase has not yet been carried out.