Preparation method and spinning of high-concentration recombinant spider silk protein spinning solution

A technology of spidroin and solution, which is applied in the field of artificial spider silk preparation, which can solve the problems of long-term, unsuitable high-molecular-weight recombined spidroin, environmental pollution, etc., and achieves improved toughness, difficulty in aggregating into disordered precipitates, and stable properties Effect

A technology of spidroin and solution, which is applied in the field of artificial spider silk preparation, which can solve the problems of long-term, unsuitable high-molecular-weight recombined spidroin, environmental pollution, etc., and achieves improved toughness, difficulty in aggregating into disordered precipitates, and stable properties Effect

CN111363022AActive Publication Date: 2020-07-03SHANGHAI JIAO TONG UNIV
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  • Preparation method and spinning of high-concentration recombinant spider silk protein spinning solution
  • Preparation method and spinning of high-concentration recombinant spider silk protein spinning solution
  • Preparation method and spinning of high-concentration recombinant spider silk protein spinning solution

Examples

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Effect test

Embodiment 1

[0037] Such as figure 1 As shown, the recombinant spidroin protein used to prepare artificial spider silk involved in this embodiment is NTD-(PR) 16 -Molecular architecture of the CTD.

[0038] This embodiment relates to the preparation method of the above-mentioned recombinant spidroin solution, comprising the following steps:

[0039] Step 1) Escherichia coli cells expressing recombinant spidroin protein were resuspended in a wall-breaking buffer solution containing 2 mol thiourea and 8 mol urea at a mass ratio of 1:10, stirred for 12 hours, and then centrifuged to collect the supernatant to obtain the recombinant Spider silk protein mixture A;

[0040] Step 2) Adjust the pH of the mixed solution A obtained in step 1 to 4.0 to precipitate some impurities, and collect the supernatant by centrifugation to obtain the recombinant spidroin protein mixed solution B;

[0041] Step 3) Dialyzing the mixed solution B into a buffer solution containing 0.3 mol of sodium chloride to o...

Embodiment 2

[0052] This example relates to high component NTD-(PR) 64 -The preparation method of CTD recombinant spidroin protein solution, comprises the following steps:

[0053] Step 1) Escherichia coli cells expressing recombinant spidroin protein were resuspended in a wall-breaking buffer solution containing 2 mol thiourea and 8 mol urea at a mass ratio of 1:10, stirred for 12 hours, and then centrifuged to collect the supernatant to obtain the recombinant Spider silk protein mixture A;

[0054] Step 2) Adjust the pH of the mixed solution A obtained in step 1 to 4.0 to precipitate some impurities, and collect the supernatant by centrifugation to obtain the recombinant spidroin protein mixed solution B;

[0055] Step 3) Dialyzing the mixed solution B into a buffer solution containing 0.3 mol of sodium chloride to obtain the reconstituted spidroin protein mixed solution C;

[0056] Step 4) Heating the mixed solution C in a water bath at 80° C. for 20 minutes to precipitate some impuri...

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Abstract

The invention discloses a preparation method and spinning of a high-concentration recombinant spider silk protein spinning solution. The molecular structure of a recombinant spider silk protein is NTD-(RP)n-CTD, an integer n represents the number of repetitions of an RP repetitive structural domain, and the amino acid sequence of the RP repetitive structural domain is shown as Seq ID No.1 or Seq ID No.2. The method can prepare the high-concentration recombinant spider silk protein spinning solution efficiently at low cost, and obtain artificial spider silks which are easy to modify and excellent in mechanical property.

Description

technical field [0001] The invention relates to a technology in the field of biopolymers, in particular to a method for purifying recombinant spidroin protein with a concentration of 50-500 mg / mL, separating and concentrating liquid-liquid phases, and a method for preparing artificial spider silk. Background technique [0002] As one of the strongest biological materials, spider silk has superior toughness than most synthetic materials due to its high strength and good ductility. Nephila clavipes traction silk is composed of two high-molecular-weight spidroin proteins, MaSp1 and MaSp2, of which more than 80% are MaSp1. Preparation of high-concentration protein spinning solution is one of the important prerequisites for biomimetic spinning of recombinant spidroin protein. [0003] In the prior art, the purified low-concentration spidroin solution is concentrated to a high concentration by centrifugation and biomimetic spinning is performed; or the purified spidroin solution ...

Claims

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Application Information

Patent Timeline
03 Jul 2020
Publication
CN111363022A
IPC
C07K14/435; C07K1/36; C07K1/34; C07K1/30; D01F4/00
CPC
C07K14/43518; D01F4/00
Inventors
夏小霞; 胡春飞