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Serial medicine carriers, tyrosine hydrozylase fusion protein as new carrier and its prepn

A fusion protein and carrier technology, applied in the field of peptide chemistry, can solve problems such as toxic side effects and curative effect decline

Inactive Publication Date: 2003-01-01
牛勃
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Oral supplementation of DA precursor levodopa (L-DOPA) is currently the most effective treatment, but generally after 3-5 years of taking the drug, the curative effect declines significantly and serious side effects occur

Method used

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  • Serial medicine carriers, tyrosine hydrozylase fusion protein as new carrier and its prepn
  • Serial medicine carriers, tyrosine hydrozylase fusion protein as new carrier and its prepn

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0012] Example 1 Preparation of PMPV gene vector:

[0013] 1). Preparation of PMPV gene:

[0014] That is, 13 lysine genes were prepared in series—entrusted to Shanghai Sangong Co., Ltd.

[0015] 2). The PMPV gene is combined with the carrier to form the PMPV gene carrier:

[0016] The vector TransVectortm is provided by Q.BIO gene company.

[0017] 1 ug of the vector TransVectortm and PMPV genes were taken, digested with type II restriction enzymes EcoRI and PstI respectively, and then recovered with a plasmid extraction kit (Wizard PCR Preps DNA Purification System). The vector and PMPV gene were mixed at a ratio of 1:3, and 5U of T was added. 4 Ligase, react at 16°C for 16 hours. CaC for reaction product l 2 BL21 bacteria were transformed by heat shock method. The transformed bacteria were plated on LB fixed plate medium containing ampicillin and X-gal for blue-white colony screening. Pick the white colony containing the recombinant and name it BL21PMPV.

[0018] P...

Embodiment 2

[0019] The total RNAlug of human fetus was taken, and each reaction component was added in sequence according to the operation guide of cDNA synthesis system. The total reaction volume was 20 ul, and after 15 min at 42 °C, it was inactivated at 99 °C for 5 min, and used as a PCR template. The total volume of the PCR reaction system is 50 ul, containing 10 ul of reverse transcription product, and the addition of other components is still carried out according to the above-mentioned operation guide. The first step: start Golden Tap enzyme at 95°C for 12min; the second step: denaturation at 94°C for 50s; the third step: anneal at 54°C for 1min; the fourth step: extend at 72°C for 1.5min; the fifth step: cycle 35 times. PCR amplification bands were checked by 2% agarose gel electrophoresis. Example 3 Preparation of PMPV-TH fusion protein gene vector ( figure 1 ):

Embodiment 3

[0019] The total RNAlug of human fetus was taken, and each reaction component was added in sequence according to the operation guide of cDNA synthesis system. The total reaction volume was 20 ul, and after 15 min at 42 °C, it was inactivated at 99 °C for 5 min, and used as a PCR template. The total volume of the PCR reaction system is 50 ul, containing 10 ul of reverse transcription product, and the addition of other components is still carried out according to the above-mentioned operation guide. The first step: start Golden Tap enzyme at 95°C for 12min; the second step: denaturation at 94°C for 50s; the third step: anneal at 54°C for 1min; the fourth step: extend at 72°C for 1.5min; the fifth step: cycle 35 times. PCR amplification bands were checked by 2% agarose gel electrophoresis. Example 3 Preparation of PMPV-TH fusion protein gene vector ( figure 1 ):

[0020] 1) Take 1ug of each of PMPV vector and TH gene, digest with type II restriction enzymes EcoRI and PstI resp...

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Abstract

The serial medicine carriers are named polypeptide mediated penetrating vector and belongs to membrane penetrating peptide family. They have the functions of penetrating cell membrane and cytoblast membrane, and may carry medicine molecule to penetrate physiological barriers, such as cell membrane, blood brain barrier, placenta barrier, etc. The new carrier-tyrosinie hydrozylase fusion protein is one new kind of protein with the features of both new carrier and tyrosinie hydrozylase, and it may reach human brain substantia nigra pathologic change part via blood brain barrier to treat parkinsonism.

Description

Technical field: [0001] The present invention relates to the field of peptide chemistry, more particularly, to a novel carrier and its preparation and use, as well as the preparation and use of a novel carrier-tyrosine hydroxylase (TH) fusion protein. Background technique: [0002] So far, most drugs cannot directly reach the lesions due to the selective permeation of human physiological barriers, especially the cell membrane, blood-brain barrier, placental barrier, blood-testis barrier, etc., which greatly reduces the amount of drugs in the body. effect, limiting the drug's treatment of the disease, affecting the efficacy. [0003] Parkinson's disease (PD) is a progressive disease of the central nervous system more common in middle-aged and elderly people. Its main pathological feature is the degeneration and necrosis of dopaminergic The DA transported to the striatum by the striatal-striatal pathway is significantly reduced, resulting in clinical symptoms such as tremor, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/16A61K47/42A61P25/16C07K14/00C07K19/00C12N15/11C12N15/62C12N15/63
Inventor 牛勃杨琦解军刘红林杨涛常冰梅张悦红郭勇阎占清
Owner 牛勃
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