Method for purifying recombinant human proinsulin

Abstract

The invention belongs to the field of biomedicine, and particularly relates to a method for purifying recombinant human proinsulin, in particular to a method for purifying recombinant human proinsulin by using macroporous absorbent resin and cation exchange resin. In the method, a recombinant human proinsuli plural liquid is adsorbed and concentrated through the macroporous absorbent resin, and is loaded onto the cation exchange resin for further purifying, so that the collected proinsulin has high yield and high purity, and can be directly applied to enzyme digestion to obtain insulin. The method is easy and convenient to operate, has high process cohesion, and is suitable for industrial production.

Description

technical field [0001] The invention belongs to the field of biomedicine, and specifically relates to a method for purifying recombinant human proinsulin, in particular to a method for purifying recombinant human proinsulin by using macroporous adsorption resin and cation exchange resin. Background technique [0002] Insulin is an irreplaceable specific drug for diabetes. Now there are more than 120 million diabetic patients in the world, and this number is increasing every year. It is estimated to be 300 million by 2025. Each diabetic patient needs to use 1.5-2.0 mg of insulin per day , the world consumes about 5,000 kilograms of insulin every year. Therefore, it is imperative to develop a low-cost method for large-scale preparation of insulin. [0003] Insulin is a protein hormone secreted by pancreatic beta cells stimulated by endogenous or exogenous substances such as glucose, lactose, ribose, arginine, glucagon, etc. It is the only hormone that lowers blood sugar in th...

AI Technical Summary

Problems solved by technology

Chinese invention patent application CN101717442A discloses the technical scheme of PEGylated recombinant human DesB30 insulin and its preparation method and application. In the technical scheme, the host cell of the recombinant expression vector is Pichia pastoris, and the purification method of proinsulin is fermentation broth After filtration, it is adsorbed by a macroporous resin and eluted with 60% ethanol, and then further purified by strong cation exchange chromatography, and then purified by zinc chloride precipitation. The proinsulin precipitate is collected and redissolved for enzyme digestion to obtain insulin. The production process for the preparation of proinsulin by this scheme is cumbersome, and proinsulin exists in the form of precipitation, which is not conducive to the direct implementation of the enzyme digestion step.

[0005] Chinese Invention Patent Specification CN88102311.6 discloses a human crystalline proinsulin and its production method. In this technical scheme, a cationic salt is first added to the solution containing human proinsulin, and then the pH is adjusted to 5.4-6.5 to form crystals, and then the crystals are recovered. Human proinsulin, although this method simplifies the production process, but the product yield is low, and proinsulin still exists in the form of precipitation

Chinese invention patent specification ZL98812757.1 discloses the preparation method of human proinsulin. The technical scheme is: the inclusion body is firstly dissolved and sulfonated, then treated with cyanogen bromide and purified by ion exchange chromatography, and then refolded. Human proinsulin is subjected to adsorption chromatography and zinc chloride precipitation to obtain human proinsulin in the form of precipitation, which can obtain a yield of 90%. The production process of this method is complicated, and the product still exists in the form of precipitation

[0006] In the method disclosed above, the operation process is complicated, and the precursor of insulin, proinsulin, finally exists in the form of precipitation, which is not conducive to the direct operation of the subsequent enzymatic digestion step, and the process connection is poor, and the yield and purity of proinsulin cannot be taken into account at the same time.