Recombinant humanized type iii collagen high expression design and industrialized preparation and application

Abstract

The present application relates to a kind of recombinant human-like source Ⅲ collagen COL3A1-6 and its preparation method and application.The present application is designed by a series of mutant recombinant human-like source Ⅲ collagen, according to the hydrophobic characteristics of mutant protein, expression, cell repair and other functions are screened, and finally obtain the excellent effect of recombinant human-like source Ⅲ collagen COL3A1-6.The product expression of the recombinant human-like source Ⅲ collagen COL3A1-6 of the present application yeast fermentation expression 72h can reach 15g / L, is three times the fermentation expression of prior art collagen COL3A1-QS-1 under the same conditions.The recombinant human-like source Ⅲ collagen COL3A1-6 of the present application has the high yield of industrial production level, and has excellent moisturizing effect, promote cell healing function, anti-wrinkle firming effect, and the corresponding freeze-dried powder preparation can be widely used in medical beauty cosmetics or functional cosmetic field.

Claims

1. A recombinant human-like type III collagen, named COL3A1-6, characterized in that, The amino acid sequence of the recombinant human type III collagen is shown in SEQ ID NO:

1.

2. An isolated nucleic acid, characterized in that, The recombinant human type III collagen of claim 1 is encoded.

3. The nucleic acid according to claim 2, characterized in that, The nucleic acid sequence is shown in SEQ ID NO:

2.

4. A recombinant vector, characterized in that, It contains the nucleic acid described in claim 2 or 3.

5. A recombinant host cell, characterized in that, It comprises the nucleic acid of claim 2 or 3, or the recombinant vector of claim 4.

6. A composition, characterized in that, It comprises the recombinant human type III collagen of claim 1, the nucleic acid of claim 2 or 3, the recombinant vector of claim 4, or the recombinant host cell of claim 5.

7. The composition according to claim 6, characterized in that, The composition is a lyophilized powder.

8. A method for preparing the recombinant human type III collagen of claim 1 by fermentation expression, characterized in that, The method includes seed culture, fermentation culture, and preparation of fermentation supernatant; wherein the seed is a seed culture of a glycerol strain of Pichia pastoris engineered strain encoding the target gene of recombinant human type III collagen as described in claim 1.

9. The method according to claim 8, characterized in that, The method includes the following steps: (1) Seed culture, wherein the seed is a seed culture of a glycerol strain of Pichia pastoris engineered strain containing the target gene encoding the recombinant human type III collagen of claim 1; (2) Fermentation culture: The seed culture is inoculated into the fermentation medium and cultured in a fermenter. The appropriate DO, stirring speed, aeration rate and temperature are controlled. Methanol is added at the appropriate growth stage to induce expression. (3) Preparation of fermentation supernatant: After culturing for a suitable time, the fermentation broth is taken to remove the cells, filtered, and fermentation supernatant is obtained.

10. A method for purifying the recombinant human type III collagen of claim 1, characterized in that, The method includes the step of ultrafiltration and concentration of the fermentation supernatant of the recombinant human type III collagen to obtain purified recombinant human type III collagen.

11. A method for preparing the lyophilized powder of the recombinant human type III collagen as described in claim 1, characterized in that, The method includes pre-lyophilizing the purified recombinant human type III collagen, and then lyophilizing it.

12. The use of the recombinant human type III collagen of claim 1, the nucleic acid of claim 2 or 3, the recombinant vector of claim 4, or the recombinant host cell of claim 5 in any of the following: (1) Preparation of functional cosmetics; (2) To prepare medical aesthetic cosmetics; or, (3) Preparation of medical dressings.

Images