Pluri-negative-charge hemoglobin alpha subunit

A technology of hemoglobin and α subunits, applied in the field of biomedicine, can solve problems such as inability to completely eliminate side effects, and achieve the effect of solving human adverse reactions

Inactive Publication Date: 2014-11-26
王子元 +2
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, these methods do not completely eliminate side effects

Method used

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  • Pluri-negative-charge hemoglobin alpha subunit
  • Pluri-negative-charge hemoglobin alpha subunit
  • Pluri-negative-charge hemoglobin alpha subunit

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Experimental program
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Effect test

Embodiment

[0027] 1. The amino acid sequence of the polypeptide chain encoding the fusion protein is synthesized by chemical total synthesis, and its DNA sequence is shown in Sequence Table 2 below.

[0028] The DNA sequence has the following amino acid sequence protein: amino acid 1-586 is human serum albumin sequence; 587-595 is connecting peptide (italics); the last 596-736 is human hemoglobin α subunit, as shown in sequence table three .

[0029] 2. Cloning the above sequence into the prokaryotic expression vector pSumo-Mut, which contains 6xHis-Sumo "tag". The restriction sites used for cloning ensure that this "tag" can be correctly added to the N-terminus of the recombinantly expressed polynegatively charged hemoglobin alpha subunit.

[0030] 3. Transforming the pSumo-Mut expression vector into Escherichia coli cells;

[0031] 4. Identify cells containing the amino acid sequence of the polypeptide chain of the fusion protein in E. coli cells by an IPTG-induced method;

[0032] ...

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Abstract

The invention relates to a pluri-negative-charge hemoglobin alpha subunit, and relates to a component of a hemoglobin red blood cell substitute artificial blood. The invention relates to the technical field of biomedicine. The pluri-negative-charge hemoglobin alpha subunit is a fusion protein composed of human serum albumin and human hemoglobin alpha subunit. The human serum albumin and human hemoglobin are connected through connecting peptide. The preparation method comprises the following steps: with a PCR amplification method or a chemical total synthesis method, a fusion protein polypeptide chain amino acid sequence is synthesized; the fusion protein polypeptide chain amino acid sequence is cloned to a prokaryotic expression vector; the expression vector is transformed into cells of an expression system; with an induction method, effective cells are identified; and the cells are subjected to separation and purification. The fusion protein has large molecular weight and carries a lot of negative charges. With the two factors, even a fusion protein in the form of a monomer cannot pass kidney filtration, such that side effect to ahuman body is reduced. The human serum albumin can be bonded with heme. Therefore, when the fusion protein is used for preparing artificial blood, higher bonding capability and higher oxygen transportation capacity are provided.

Description

technical field [0001] The invention relates to a component of hemoglobin erythrocyte substitute artificial blood: multiple negatively charged hemoglobin α subunits, belonging to the technical field of biomedicine. Background technique [0002] Blood transfusion is a necessary basic treatment in clinical treatment. As the most populous country in the world, China has a large number of trauma, surgery and emergency patients who need blood transfusion every day. Especially in the scene of various natural disasters, major public health emergencies and military conflicts, the demand for blood will increase significantly, and artificial blood that is easy to preserve and use is of great significance for saving lives. In addition, my country will gradually enter an aging society, and the contradiction between blood supply and demand will become more prominent. At present, the quantity or quality of blood products in our country is insufficient or not high enough to meet clinical...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/70
Inventor 王子元许铁刘抗王菲傅学斌燕宪亮杨红宁
Owner 王子元
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