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Human immunoglobulin-like receptor mutant and application thereof

A technology for human immunoglobulins and mutants, applied in animal/human proteins, receptors/cell surface antigens/cell surface determinants, specific peptides, etc., can solve the problem of low ILT3 efficiency, inability to adapt to preservation, ILT3 extracellular domain Problems such as protein instability, to achieve the effect of improving stability and preparation efficiency, significant economic value and social significance

Inactive Publication Date: 2010-09-15
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the efficiency of purifying ILT3 in vitro by conventional methods is low, the phenomenon of protein denaturation and precipitation is serious, and the obtained extracellular domain protein of ILT3 is also unstable and unable to adapt to long-term storage, which restricts the research on its function and the utilization of its potential value.

Method used

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  • Human immunoglobulin-like receptor mutant and application thereof
  • Human immunoglobulin-like receptor mutant and application thereof
  • Human immunoglobulin-like receptor mutant and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1, Preparation of Human Immunoglobulin-like Receptor Mutants

[0037] 1. Bioinformatics analysis

[0038] The analysis found that the flexible loop region of the Ig domain 2 of the ILT3 protein had an adverse effect on the structural rigidity of the entire protein. Sequence comparison analysis of ILT 2 / 4 / 11, a member of the ILT family, found that they have a common feature, that is, each immunoglobulin domain is composed of two disulfide bonds (disulfide bond 1 and disulfide bond 2) Fixed to form a stable folding mode, the two disulfide bonds are formed by two pairs of highly conserved cysteines. For members of ILT 2 / 3 / 4 / 11, except for two pairs of highly conserved cysteines, other cysteines are non-conserved. As long as the non-conserved cysteine ​​exists, it always appears in pairs to form a disulfide bond (disulfide bond 3), and the position of the disulfide bond is fixed. ILT3 and ILT11 do not have this non-conserved disulfide bond, which makes the loop r...

Embodiment 2

[0074] Example 2, Determination of Physiological and Biochemical Properties of ILT3cc Protein

[0075] The purified ILT3cc protein and ILT3wt protein (control) prepared in Example 1 were identified as follows:

[0076] 1. Molecular sieve chromatography

[0077] ILT3cc protein and ILT3wt protein were purified with Superdex 75HR molecular sieve chromatography column (GE Healthcare) and concentrated. Molecular sieve chromatography see image 3 , the results showed that the properties of ILT3cc and ILT3wt solutions remained consistent.

[0078] After molecular sieve chromatography, the collected ILT3cc protein was subjected to SDS-PAGE, and the electropherogram is shown in Figure 4 . Place the ILT3cc protein and ILT3wt protein at 4°C for 24 hours, and then perform electrophoresis. The results are shown in Figure 5 . The results showed that after standing for 24 hours, compared with ILT3wt protein, the degradation of ILT3cc protein was significantly improved.

[0079] 2. P...

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Abstract

The invention discloses a human immunoglobulin-like receptor mutant and application thereof. The human immunoglobulin-like receptor mutant provided by the invention is obtained by introducing a disulfide bond into a human immunoglobulin-like receptor. The invention also provides a method for improving the stability of the human immunoglobulin-like receptor, which is to introduce the disulfide bond into the human immunoglobulin-like receptor. The human immunoglobulin-like receptor mutant of the invention can be used in the structural analysis of the human immunoglobulin-like receptor and can also be used in researches on organ transplanting and tumor transfer and the screening of medicinal targets. The mutant and the application thereof have a high economic value and a great social significance.

Description

technical field [0001] The invention relates to a human immunoglobulin-like receptor mutant and its application. Background technique [0002] Organ transplantation is the ultimate effective way to save the lives of patients with end-stage organ failure. How to control the rejection response after organ transplantation and induce the host's immune tolerance to the graft has always been a hot spot and difficult problem in the field of transplantation immunology. So far, the main countermeasure to prevent and treat transplant rejection is still to suppress the immune function of the host through various channels. It is also due to the advent of immunosuppressive drugs such as cyclosporine, FK506 and OKT3, including kidney, liver, pancreas and skin, etc. However, the non-specificity of its inhibitory effect has not been well resolved. The theory of specific immune tolerance proposed in recent years has pointed out the direction for the research of transplant immunity: that is...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/705
Inventor 高福陈勇
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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