CN106749629A - A kind of method of collagen self assembly - Google Patents
A kind of method of collagen self assembly Download PDFInfo
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- CN106749629A CN106749629A CN201611153878.4A CN201611153878A CN106749629A CN 106749629 A CN106749629 A CN 106749629A CN 201611153878 A CN201611153878 A CN 201611153878A CN 106749629 A CN106749629 A CN 106749629A
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- CN
- China
- Prior art keywords
- collagen
- self assembly
- infrared radiation
- polyethylene glycol
- constant temperature
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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- 102000008186 Collagen Human genes 0.000 title claims abstract description 61
- 108010035532 Collagen Proteins 0.000 title claims abstract description 61
- 229920001436 collagen Polymers 0.000 title claims abstract description 61
- 238000001338 self-assembly Methods 0.000 title claims abstract description 53
- 238000000034 method Methods 0.000 title claims abstract description 25
- 239000002202 Polyethylene glycol Substances 0.000 claims abstract description 16
- 229920001223 polyethylene glycol Polymers 0.000 claims abstract description 16
- 230000005855 radiation Effects 0.000 claims abstract description 15
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 24
- 230000020978 protein processing Effects 0.000 abstract description 2
- 238000001514 detection method Methods 0.000 description 5
- PMMYEEVYMWASQN-DMTCNVIQSA-N Hydroxyproline Chemical class O[C@H]1CN[C@H](C(O)=O)C1 PMMYEEVYMWASQN-DMTCNVIQSA-N 0.000 description 3
- PMMYEEVYMWASQN-UHFFFAOYSA-N dl-hydroxyproline Natural products OC1C[NH2+]C(C([O-])=O)C1 PMMYEEVYMWASQN-UHFFFAOYSA-N 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 229960002591 hydroxyproline Drugs 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- FGMPLJWBKKVCDB-UHFFFAOYSA-N trans-L-hydroxy-proline Natural products ON1CCCC1C(O)=O FGMPLJWBKKVCDB-UHFFFAOYSA-N 0.000 description 2
- DGPBVJWCIDNDPN-UHFFFAOYSA-N 2-(dimethylamino)benzaldehyde Chemical compound CN(C)C1=CC=CC=C1C=O DGPBVJWCIDNDPN-UHFFFAOYSA-N 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004166 bioassay Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 230000007261 regionalization Effects 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/78—Connective tissue peptides, e.g. collagen, elastin, laminin, fibronectin, vitronectin or cold insoluble globulin [CIG]
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- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Gastroenterology & Hepatology (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Toxicology (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
The invention discloses a kind of method of collagen self assembly, belong to protein processing technique field.The present invention carries out self assembly using the collagen that molecular weight is 250 350kDa, adds polyethylene glycol, and regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake self assembly 3 6h.The concentration of collagen is 6 8mg/mL.The addition of polyethylene glycol is the 2 5% of collagen solution volume.The 400W of infrared radiation processing power 200 described in infrared radiation, 40 60 μm of wave-length coverage.The collagen self-assembling method that the present invention is provided, simple and fast significantly improves collagen self assembly degree, shortens the autonomous ETL estimated time of loading of collagen, the self assembly degree of collagen is improved to 93%, and built-up time foreshortens to 3 6h.
Description
Technical field
Patent of the present invention is related to a kind of method of collagen self assembly, belongs to protein processing technique field.
Background technology
Molecular self-assembling phenomenon generally existing in living nature, the self assembly of macromolecular complex or macromolecular are big with other
Self assembly between molecule can form the biological structure of various complexity.Specific self assembling process is due to non-covalent bond
Effect, the molecule self-assemble in system is assembled into the structure with certain rule, and molecular self-assembling is not a large amount of molecule letters
It is singly added together, but a kind of sufficiently complex overall coordination effect.
The complex structure of collagen, very big difficulty is brought to research collagen assembly study mechanism, and research finds, biological
It is notable at aspects such as heat endurance, resistance to enzyme degradability and biomechanical properties with the collagenous fibres of self assembly pattern formation in vivo
Better than the external performance of collagen monomer molecule, but due to the study mechanism also imperfection of self assembly, collagen from
A still technical problem for being difficult to capture is assembled, often there is a problem of can't detect assembly or self assembly low degree.
The content of the invention
It is an object of the invention to provide a kind of method of collagen self assembly, solve in the prior art collagen from group
Fill the technical problem of low degree.
A kind of method of collagen self assembly, using molecular weight for the collagen of 250-350kDa carries out self assembly,
Polyethylene glycol is added, regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake self assembly 3-
6h。
The concentration of the collagen is 6-8mg/mL.
The addition of the polyethylene glycol is the 2-5% of collagen solution volume.
Infrared radiation processing power 200-400W described in the infrared radiation, 40-60 μm of wave-length coverage.
The constant temperature is 30-35 DEG C of temperature.
Shaking table shake speed is 10r/min.
With the NaOH regulation system pH value of 0.1mol/L.
The beneficial effects of the present invention are：
(1) the collagen self-assembling method that the present invention is provided, simple and fast significantly improves collagen self assembly journey
Degree, shortens the autonomous ETL estimated time of loading of collagen, and the self assembly degree of collagen is improved to 93%, and built-up time foreshortens to 3-
6h。
Specific embodiment
Self assembly level determinations
With L- hydroxyprolines as reference standards, surveyed using spectrophotometric standard measure by developer of dimethylaminobenzaldehyde
Determine Hydroxyproline content in centrifuged supernatant, bioassay standard curve is：Y=0.19301x+0.01521, R2=0.9997, formula
In：Y is hydroxyproline content/(mg/mL), and x is mensuration absorbance.Collagen assembly degree is calculated by formula (1).
In formula：m1It is the THP/mg in assembling precollagen sample；m2It is the hydroxyl after assembling in sample supernatant
Proline total amount/mg.
Embodiment 1
A kind of method of collagen self assembly, is that the upper and lower collagens of 300kDa carry out self assembly using molecular weight,
Regulation pH value is 8.0, the self assembly 4h under constant temperature.The concentration of the collagen is 7mg/mL.The polyethylene glycol
Addition is the 4% of collagen solution volume.The constant temperature is 30 DEG C of temperature.With the NaOH regulation systems of 0.1mol/L
PH value.After 4h assemblings are finished, detection self assembly degree is 45%.
Embodiment 2
A kind of method of collagen self assembly, is that the upper and lower collagens of 300kDa carry out self assembly using molecular weight,
Regulation pH value is 8.0, the self assembly 16h under constant temperature.The concentration of the collagen is 7mg/mL.The polyethylene glycol
Addition is the 4% of collagen solution volume.The constant temperature is 30 DEG C of temperature.With the NaOH regulation systems of 0.1mol/L
PH value.After 4h assemblings are finished, detection self assembly degree is 82%.
Embodiment 3
A kind of method of collagen self assembly, is that the upper and lower collagens of 300kDa carry out self assembly using molecular weight,
Polyethylene glycol is added, regulation pH value is 8.0, under constant temperature, using shaking table shake self assembly 4h.The collagen it is dense
It is 7mg/mL to spend.The addition of the polyethylene glycol is the 4% of collagen solution volume.The constant temperature is temperature 30
℃.Shaking table shake speed is 10r/min.With the NaOH regulation system pH value of 0.1mol/L.After 4h assemblings are finished, self assembly is detected
Degree is 64%.
Embodiment 4
A kind of method of collagen self assembly, is that the upper and lower collagens of 300kDa carry out self assembly using molecular weight,
Polyethylene glycol is added, regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake self assembly 4h.
The concentration of the collagen is 7mg/mL.The addition of the polyethylene glycol is the 3% of collagen solution volume.It is described red
Infrared radiation processing power 300W described in external irradiation, 40 μm of wave-length coverage.The constant temperature is 30 DEG C of temperature.Shaking table shake speed
It is 10r/min to spend.With the NaOH regulation system pH value of 0.1mol/L.After 4h assemblings are finished, detection self assembly degree is 87%
Embodiment 5
A kind of method of collagen self assembly, is that the upper and lower collagens of 350kDa carry out self assembly using molecular weight,
Polyethylene glycol is added, regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake self assembly 6h.
The concentration of the collagen is 8mg/mL.The addition of the polyethylene glycol is the 5% of collagen solution volume.It is described red
Infrared radiation processing power 400W described in external irradiation, 40 μm of wave-length coverage.The constant temperature is 35 DEG C of temperature.Shaking table shake speed
It is 10r/min to spend.With the NaOH regulation system pH value of 0.1mol/L.After 6h assemblings are finished, detection self assembly degree is 93%.
Embodiment 6
A kind of method of collagen self assembly, is that the upper and lower collagens of 60kDa carry out self assembly using molecular weight, plus
Enter polyethylene glycol, regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake self assembly 6h.Institute
The concentration for stating collagen is 8mg/mL.The addition of the polyethylene glycol is the 5% of collagen solution volume.It is described infrared
Irradiate the infrared radiation processing power 400W, 40 μm of wave-length coverage.The constant temperature is 35 DEG C of temperature.Shaking table shakes speed
It is 10r/min.With the NaOH regulation system pH value of 0.1mol/L.After 6h assemblings are finished, detection self assembly degree is 44%.
Claims (8)
1. a kind of method of collagen self assembly, it is characterised in that using molecular weight for the collagen of 250-350kDa enters
Row self assembly, adds polyethylene glycol, and regulation pH value is 8.0, under constant temperature, using infrared radiation, and using shaking table shake
Self assembly 3-6h.
2. the method for a kind of collagen self assembly according to claim 1, it is characterised in that the collagen it is dense
It is 6-8mg/mL to spend.
3. the method for a kind of collagen self assembly according to claim 1, it is characterised in that the polyethylene glycol adds
Dosage is the 2-5% of collagen solution volume.
4. the method for a kind of collagen self assembly according to claim 1, it is characterised in that described in the infrared radiation
Infrared radiation processing power 200-400W, 40-60 μm of wave-length coverage.
5. the method for a kind of collagen self assembly according to claim 1, it is characterised in that the constant temperature is temperature
30-35 DEG C of degree.
6. the method for a kind of collagen self assembly according to claim 1, it is characterised in that shaking table shake speed is
10r/min。
7. the method for a kind of collagen self assembly according to claim 1, it is characterised in that with the NaOH of 0.1mol/L
Regulation system pH value.
8. the collagen assembly for being prepared according to any methods describeds of claim 1-7.
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|---|---|---|---|
| CN201611153878.4A CN106749629A (en) | 2016-12-14 | 2016-12-14 | A kind of method of collagen self assembly |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201611153878.4A CN106749629A (en) | 2016-12-14 | 2016-12-14 | A kind of method of collagen self assembly |
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| CN201611153878.4A Pending CN106749629A (en) | 2016-12-14 | 2016-12-14 | A kind of method of collagen self assembly |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113773379A (en) * | 2021-09-13 | 2021-12-10 | 熹微(苏州)生物医药科技有限公司 | Method for preparing polyethylene glycol collagen and application thereof |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105802252A (en) * | 2016-04-29 | 2016-07-27 | 苏州景卓生物技术有限公司 | Collagen modifying method and modified collagen prepared by same |
-
2016
- 2016-12-14 CN CN201611153878.4A patent/CN106749629A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105802252A (en) * | 2016-04-29 | 2016-07-27 | 苏州景卓生物技术有限公司 | Collagen modifying method and modified collagen prepared by same |
Non-Patent Citations (1)
| Title |
|---|
| 闫婷婷等: "胶原大分子自组装研究进展", 《高分子通报》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113773379A (en) * | 2021-09-13 | 2021-12-10 | 熹微(苏州)生物医药科技有限公司 | Method for preparing polyethylene glycol collagen and application thereof |
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