Protein analysis: saturation mutation and continuous evolution

Posted by beauty33 on May 23rd, 2019

Protein dialysis is an important part of protein-related research. After cell lysis or separation, lysates often contain pollutants incompatible with downstream steps. A variety of devices and resins are provided in the preparation process for simple and efficient desalination, buffer replacement, and removal of scale agents from samples. In addition, if the protein concentration is too low for subsequent experiments and analysis, you can use a centrifuge concentrator to rapidly concentrate the sample. At the same time, in order to solve the defects of various resins and equipment, it is necessary to carry out effective and rapid desalting and detergent removal for samples. Of course, the use of natural protein as a preparation can effectively solve the above problems,natural protein requires the use of special tools to obtain, the amino acid composition, size, gene sequence and other characteristics of this protein has experimental advantages. Protein saturation mutagenesis can generate random mutations, and the field of protein evolution and engineering research currently has advanced phage assisted continuous evolution services.

Protein liposome dialysis separation technique based on semi - dialysis

Proteoliposome dialysis separation technology is a kind of separation and purification technology to separate small molecules and biomacromolecules by using the principle of small molecules diffusing to liquid through the semi-permeable membrane. At present, dialysis membrane is often used in medical field, such as hemodialysis and peritoneal dialysis, used in the treatment of acute or chronic renal failure, drug or other chemicals in the body accumulation, stay in the level of health requires high separation products, such as pharmaceutical products, cosmetics products, also can use a semipermeable membrane dialysis to separate products. Due to the use of a semipermeable membrane separation device of dialysis method is mostly used in medical field, with a semipermeable membrane dialysis dialysis by a semipermeable membrane separation device generally set up the adapted with dialysis for a semipermeable membrane separation in the space, but for the dialysis membrane size limit, make each time through a semipermeable membrane dialysis after separation of the finished product quantity is limited, a semipermeable membrane dialysis efficiency of separation of the ground, not suitable for industrial scale applications, and because the liquid to replace dialysis use, keep dialysis, dialysis environment and conditions of the space, making a semi-permeable membrane dialysis separation device operation is quite complicated.

Protein liposomes

Liposome protein in organism can degrade nontoxic and no immunogenicity, as drug carrier, has targeted, thus reducing drug dose, lower toxicity, decrease side effects, etc. Compared with the traditional liposomes, compound emulsion prepared multivesicular liposomes, also has a discontinuous vesicle drug solution, the vesicles were continuous excentrically lipid double molecular lecithin membrane separation, has more coating volume and larger size, when some vesicles rupture, vesicle release drugs only from burst, complete vesicles can still maintain the original state, thus has good slow release effect. But it is also the same as other traditional liposomes, poor stability and easy Liposomes are passive collectors of drugs, so low encapsulation rate is also a difficult problem for water-soluble proteins. After digestion, a large number of unencapsulated protein drugs will be lost, and the separation of liposomes is a complicated process.

Protein saturation mutagenesis

Point saturation mutation technology is a new technology in protein engineering. It obtains mutants whose target amino acids are replaced by 19 other amino acids in a short time by modifying the coding genes of the target protein. This technique is not only a powerful tool for directed protein modification, but also an important means for studying the relationship between protein structure and function. Site saturation mutagenesis (SSM) is a PCR amplification method using degenerate synthesis of oligonucleotides as protein primers. Because specific amino acid residues in protein sequences can be identified, it is important to determine the ideal amino acid residues for the position of oligonucleotides.

Phage - assisted continuous directional polymorphic evolution system and method

Phage-assisted multi-bacterial continuous directed evolution system and method. This system contains phage SM carrying the target gene to be evolved, supporting multiple helper plasmid HP proliferating in different SM before and after evolution and plasmid IP inducing mutation. Different HP can be placed in different host bacteria, which can effectively improve the proliferation and evolution efficiency of phage and avoid the mutual interference between different gene elements. Therefore, it has good practicability and can be used for the directional evolution of multiple genes. In addition, because phage cells that are required to express phage genes are infectious in vivo, they can continue to infect and reproduce in fresh host cells diluted in blood vessels, which will play an important role in protein-saturated mutation and continuous evolution.