CN106554968A - Schistosoma japonicum recombiant protein and its preparation method and application - Google Patents
Schistosoma japonicum recombiant protein and its preparation method and application Download PDFInfo
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Abstract
The invention discloses a kind of Schistosoma japonicum recombiant protein, the recombiant protein is obtained by the recombinant vector Jing expression of the genes of C1qBP containing Schistosoma japonicum.The invention also discloses the preparation method of the Schistosoma japonicum recombiant protein rSjC1qBP and its application in the medicine for preparing treatment schistosomicide.The Schistosoma japonicum recombiant protein rSjC1qBP of the present invention, can induce in mouse immune experiment in mice body and produce the specific IgG antibodies of anti-recombiant protein and a higher level can be reached, show that the recombiant protein has the potentiality for developing into anti-schistosomiasis novel drugs, with good using value.The recombiant protein rSjC1qBP of the present invention can suppress complement hemolysis, and haemolysis suppression ratio shows recombiant protein rSjC1qBP of the present invention with the ability combined with complement system C1q molecule, for having a extensive future for treatment complement activity abnormal diseases up to more than 30%.
Description
Technical field
The present invention relates to technical field of bioengineering, more particularly to a kind of Schistosoma japonicum recombiant protein and its preparation method and application.
Background technology
Schistosomiasis japanica is a kind of widely distributed, the infecting both domestic animals and human parasitic disease that harm is serious caused by Schistosoma japonicum.Day
The unique intermediate host oncomelania distribution area of japonicum is wide, breeding environment is complicated, it is difficult to eliminate；Furthermore, people and animals' repeated infection
Seriously.Therefore, the schistosomiasis japanica preventing and controlling of China remain that one long-term, arduous and the task of complexity.
Schistosomulum pierces final host's skin and is changed into virgin worm, and the development after lungs, liver of dividing a word with a hyphen at the end of a line is adult, and in place
Breeding spawning, long-term parasitism in main body.The blood environment that schistosomicide is lived away from home be provided for suitable Physiology and biochemistry environment and nutrition,
The materials such as hormone, signaling molecule, but it is directly exposed in the immunological effect factor of host, such as host's complement.Complement system
System is the important component part of immunity of organism defense mechanism, and the performance of its effect relies on the activation of complement system.Complement system can be fast
Fast labelling simultaneously removes invading micro-organism or toxic component, humoral immunity and cellular immunization, is important immunomodulating in body
With effect amplification system.Schistosomulum is just attacked by host immune response after breaking through skin barrier, and complementary reaction is host
Early start kills the immunne response of polypide, is one of mechanism that host obtains non-specific drag.But schistosomicide is long-term
It is also accumulated from tackling the adaptability of host's complement system in evolutionary process, the schistosomicide of significant proportion can be escaped the immunity of host and attack
Hitting, spawning being survived, reach maturity and breed in the host blood vessel for setting up immunne response, the regulation to complementary reaction is which is exempted from
The important component part of epidemic disease avoidance strategy.Starting molecules of the C1q as classical pathway of complement, is the weight of complement system classical pathway
Molecule to be recognized, can start classical pathway, and main interconnection function is played between inherent immunity and specific immunity.
C1qBP can be combined with C1q, suppress complement activation pathway.Research SjC1qBP contributes to explanation schistosomicide immune evasion phenomenon,
The interphase interaction of polypide and host is disclosed, and C1qBP molecules is assessed as the potentiality of schistosomicide novel drugs, and to screen anti-blood
Trematodiasiss novel drugs molecule provides foundation.But, up to the present, also do not occur with regard to Schistosoma japonicum SjC1qBP restructuring eggs
The open report of Bai Zuowei antischistosomal drugs.
Complement (complement, C) is the important effect system of body inherent immunity (innate immunity), because being initial
The heat labile protein being found in serum, can aid in or " supplement " specific antibody bactericidal action and gain the name.As in vivo most
Complicated limited proteolysises system, complement are located at a line of body defenses.
Complement system is the important component part of immunity of organism defense mechanism.The activation of complement cascade reaction includes three approach, point
Wei not classical pathway, alternative route and lectin pathway.Classical pathway played a role in the effective stage of specific immunity；Substitute
Approach and alternative pathway participate in nonspecific immunity, play a significant role in infection early stage opposing cause pathogeny imcrobe infection.Complement
Reaction is very fast, Cascaded amplification immune defence mechanism, is that host kills the entozoal important channel of blood.Research table
Bright, complementary reaction is the important effect mechanism that host kills schizotrypanum cruzi.Schistosomulum is just subject to host after breaking through skin barrier
The attack of immunne response, complementary reaction are the immunne response that host's early start kills polypide, are that host is acquired non-specific anti-
One of mechanism of power.But schistosomicide is also accumulated from the adaptability for tackling host's complement system in long-term evolutionary process, to mending
The regulation of precursor reactant is the important component part of its immune evasion strategy.Schistosomicide is very sensitive to SC in the cercaria stage,
But the sensitivity that SC is killed is begun to be gradually reduced, this shows that schistosomicide exists after cercaria is changed into virgin worm 2-4 hours
Initial infection just has adjustment effect to host's complementary reaction.
The content of the invention
The invention solves the problems that lacking the technical problem of schistosomicide novel drugs at present, there is provided a kind of Schistosoma japonicum recombiant protein, should
Aminoacid sequence shown in SEQ ID NO.1 of the recombiant protein comprising Schistosoma japonicum C1qBP (SjC1qBP), with good
Immunogenicity, is adapted as anti-schistosomiasis novel drugs.
In addition, it is also desirable to provide a kind of preparation method and application of above-mentioned Schistosoma japonicum recombiant protein.
In order to solve above-mentioned technical problem, the present invention is achieved through the following technical solutions:
In one aspect of the invention, there is provided a kind of recombinant vector, comprising Schistosoma japonicum C1qBP genes, the gene order
It is the nucleotide sequence for encoding aminoacid sequence shown in SEQ ID NO.1.
Preferably, the gene order is nucleotide sequence shown in SEQ ID NO.2.
It is furthermore preferred that the empty carrier of the recombinant vector is prokaryotic expression carrier pET-32a (+), the Schistosoma japonicum C1qBP
Gene is inserted between I two restriction enzyme sites of EcoR I and Xho of empty carrier pET-32a (+).
In another aspect of this invention, a kind of Schistosoma japonicum recombiant protein is additionally provided, the recombiant protein is carried by above-mentioned restructuring
Body Jing is expressed and is obtained.
The Schistosoma japonicum recombiant protein of the present invention, also comprising being expressed by partial vector sequences with sequence labels such as His, Trx,
The sequence label is only easy for the purification of follow-up recombiant protein.
In another aspect of this invention, additionally provide a kind of host cell comprising above-mentioned recombinant vector.
In another aspect of this invention, a kind of preparation method of above-mentioned Schistosoma japonicum recombiant protein, including following step are additionally provided
Suddenly：
Build the recombinant expression carrier of the genes of C1qBP containing Schistosoma japonicum；
The recombinant expression carrier is transformed into into e. coli host cell；
E. coli host cell of the culture comprising the recombinant expression carrier, and under suitable conditions, induce the recombinant expressed load
Body surface Dagri japonicum C1qBP albumen.
Preferably, also including step：The Schistosoma japonicum C1qBP albumen that abduction delivering is obtained is further purified.
In one particular embodiment of the present invention, the recombinant expression carrier of the genes of C1qBP containing Schistosoma japonicum is built, it is concrete to wrap
Include following steps：On-line analyses are carried out using bioinformatics, the nucleotide fragments of coding Schistosoma japonicum C1qBP are found out；Root
PCR primer is designed according to the nucleotide sequence, and add specific restriction endonuclease digestion site at two ends, PCR amplification of nucleic acid segments,
Recycle specific restriction restriction endonuclease EcoR I, XhoI to encode the nucleic acid fragment directed cloning of SjC1qBP antigens to protokaryon table
Up to the multiple clone site area of carrier pET-32a (+), recombined pronucleus expression plasmid pET-32a (+)-SjC1qBP is built.
In another aspect of this invention, a kind of complement inhibitor comprising above-mentioned Schistosoma japonicum recombiant protein, the benefit are additionally provided
Body inhibitor can be applicable to biological experiment.
In another aspect of this invention, additionally provide a kind of antibody that can be specifically bound with above-mentioned Schistosoma japonicum recombiant protein.
In another aspect of this invention, additionally provide a kind of above-mentioned Schistosoma japonicum recombiant protein and prepare treatment schistosomicide medicine
In application.
In another aspect of this invention, additionally provide a kind of above-mentioned Schistosoma japonicum recombiant protein and prepare treatment complement abnormality disease
Medicine in application.
Schistosoma japonicum recombiant protein of the present invention, can induce in mice body in mouse immune experiment and produces the special of anti-recombiant protein
Property IgG antibody and a higher level can be reached, Western-blot analysis shows rSjC1qBP have preferable immunogenicity,
Show that the recombiant protein has the potentiality for developing into anti-schistosomiasis novel drugs, with good using value.And the weight of the present invention
Histone rSjC1qBP can suppress the haemolysis suppression ratio of complement hemolysis, generation to show recombiant protein of the present invention up to more than 30%
RSjC1qBP with the ability combined with complement system C1q molecule, for treatment complement activity abnormal diseases application prospect very
It is wide.
Description of the drawings
The present invention is further detailed explanation with reference to the accompanying drawings and detailed description.
Fig. 1 be the SjC1qBP of the embodiment of the present invention 1 PCR amplifications and recombined pronucleus expression plasmid pET-32a (+)-
The double digestion qualification figure of SjC1qBP；
Fig. 2 is 1 recombiant protein SjC1qBP Expression temporal analysis charts of the embodiment of the present invention；
Fig. 3 is 1 recombiant protein SjC1qBP of embodiment of the present invention expression and purification result figure；
Fig. 4 is 2 recombiant protein SjC1qBP antigenicity analysis result figures of the embodiment of the present invention；
Fig. 5 is the testing result figure of 3 anti-rSjC1qBP specific IgG antibodies level of the embodiment of the present invention；
Fig. 6 is the result figure of 4 hemolytic test of the embodiment of the present invention；
Fig. 7 is 4 Evaluation of Complement Fixation Test figure of the embodiment of the present invention.
Specific embodiment
In the following example, the experimental technique of unreceipted actual conditions, generally routinely condition, such as《Fine works molecular biology reality
Test guide》(F.M. Ao Sibai, R.E. James Kingston, J.G. Sai Deman etc. are edited, and Ma Xuejun, Su Yuelong's are translated. Beijing：Section
Learn publishing house, 2004) described in method carry out.
The present invention has cloned Schistosoma japonicum SjC1qBP (abbreviation SjC1BP) gene coded sequence using PCR, and carries out
Bioinformatic analysis, SjC1BP ORF contain 729 nucleotide, encode 242 aminoacid, and theoretical molecular is 27.58kD,
Theoretical isoelectric point, IP is 4.728；Using the transmembrane region of TMHMM software prediction protein, as a result show SjC1BP do not exist across
Film area, SignalP software predictions show that SjC1BP does not have signal peptide；The secondary structure prediction of the aminoacid shows, the gene
The polypeptide of coding based on random coil, SMART software (http://smart.embl-heidelberg.de/) its domain is observed,
Predict its complement system C1q binding site, thus it is speculated that SjC1BP molecules there may be the function of suppressing and adjust host's complementary reaction,
And important immune evasion function is played in schistosomicide divides a word with a hyphen at the end of a line growth course.
The present invention expands the nucleotide fragments for obtaining coding Schistosoma japonicum SjC1BP genes using round pcr, and uses gene
Engineering recombinant technique recombinates the nucleotide fragments Dao carrier pET-32a (+) in, constructs pET-32a (+)-SjC1BP protokaryon tables
Up to plasmid.Induction table will be carried out after recombined pronucleus expression plasmid pET-32a (+)-SjC1BP conversions e. coli bl21 (DE3)
Recombiant protein is obtained up to simultaneously purification.
After the present invention applies bioinformatics method and technique for gene engineering to obtain recombiant protein rSjC1BP, by western blot
The antigenicity and immunogenicity of this albumen are analyzed, result of the test shows that recombiant protein rSjC1BP has good antigenicity and immunity
Originality, and the specific IgG antibodies that the anti-recombinant antigen is produced in mice body, and these are can induce in mouse immune experiment
Antibody can reach a higher level.The result of its biological function research shows that recombiant protein rSjC1BP can suppress to mend
Body haemolysis, the haemolysis suppression ratio of generation up to more than 30%, with the ability combined with the C1q molecules in complement system.These
Test result indicate that, recombiant protein rSjC1qBP of the present invention can induce the high-caliber specific antibody of generation, indicate the recombiant protein
With the potentiality for developing into anti-schistosomiasis novel drugs, at the same the recombiant protein can by with reference to C1q molecules in inhibiting complement hemolysises,
Using value is wide.
The expression of 1 Schistosoma japonicum recombiant protein of embodiment and purification
1. method
The structure of 1.1 recombinant expression plasmids
The gene order (FN315565) of the SjC1qBP (abbreviation SjC1BP) included according to NCBI, designs primer, upstream
Primer (P1)：5 '-CGCGAATTCTTACGGCGTGAAATGTCT-3 ' (SEQ ID NO.3) band EcoR I enzyme action position
Point；Downstream primer (P2)：5 '-GCGCTCGAGCAGTCTTTAAGGATTGTGCA-3 ' (SEQ ID NO.4) band
Xho I restriction enzyme sites, for expanding SjC1qBP genetic fragments.CDNA with 42 days polypides of Schistosoma japonicum as template, PCR
Its cDNA fragment for containing ORF is expanded, reaction system composition is as follows：
PCR reaction conditions：94 DEG C of 5min, 94 DEG C of degeneration 30sec, 56 DEG C of annealing 30sec, 72 DEG C of extension 90sec, totally 30 are followed
Ring；Last 72 DEG C of 15min.PCR primer connects pMD19-T carriers after purification, converts DH5 α cells, picking monoclonal,
Enter the identification of performing PCR bacterium solution, positive colony is served the sequencing of Hai Huajin Bioisystech Co., Ltd, sequencing result is above carried with NCBI
The sequence of friendship is compared, and is transferred LB fluid mediums with the correct pMD19-T-SjC1qBP/DH5 α of above-mentioned sequencing, according to
Kit specification, extracts in a small amount plasmid DNA, using EcoR I, I double digestion systems of Xho in 37 DEG C of thermostats double enzymes
Cut 30min.Digestion products obtain the DNA purpose fragments with sticky end using the method for gel purification, and by its
, in the multiple clone site of prokaryotic expression carrier pET-32a (+), 16 DEG C of connections are overnight for directed cloning.Thus, build and recombinated
Plasmid pET-32a (+)-SjC1qBP, is transformed in BL21 cells, picking monoclonal, is entered the identification of performing PCR bacterium solution, is taken out
Carrying positive colony plasmid carries out double digestion identification, and serves the sequencing of Hai Huajin companies.
The expression and purification of 1.2 recombiant protein pET-32a (+)-SjC1qBP
Correct pET-32a (+)-SjC1qBP/BL21 bacterium solutions will be sequenced it be transferred to 500ml and will contain ampicillin (100 μ g/ml)
LB fluid mediums in, 37 DEG C of constant-temperature shaking cultures work as OD600When=0.6, IPTG is added so as to final concentration of 1mmol/L
Carry out abduction delivering.Take out within 0,1,2,3,4,5,6 hours after induction 1ml bacterium solutions, 4 DEG C of 12000 × g centrifugations
3min, outwells supernatant, is fully suspended precipitation with 50 μ l PBS, is subsequently added 50 μ l protein electrophoresises sample-loading buffers and blows and beats mixed
It is even.Boiling water boiling sample 5min, carries out SDS-PAGE electrophoresis.
The remaining bacterium solution induction of 6 hours, 4 DEG C of 12000 × g are centrifuged 15min, outwell supernatant, abundant with 18ml PBS
Suspend precipitation, and after multigelation three times, ice-bath ultrasonic 15min (ultrasonic 5s, stop 45s), subsequent 4 DEG C of 12000 × g are centrifuged 15min,
Collect supernatant.10ml 8mol/L carbamide dissolution precipitations are used again, and after being completely dissolved, 4 DEG C of 12000 × g are centrifuged 15min, in collection
Clearly.The Supernatant samples that will be collected, mixed protein sample-loading buffer, after boiling water boiling sample 5min, directly carry out SDS-PAGE point
Analysis.Recombiant protein successful expression in escherichia coli, it is with gained supernatant after carbamide dissolving centrifugation as sample, sharp to specifications
Purified with Ni-NTA His-Bind Resin and obtain recombiant protein.
2. result
The structure of 2.1 recombiant plasmid pET-32a (+)-SjC1qBP and identification
The SjC1qBP specific fragments (Fig. 1, left figure A swimming lane) that size is 729bp are obtained using PCR amplifications, with expection
Purpose fragment size is consistent.Using PCR and restriction endonuclease sites EcoRI, XhoI by the nucleotide fragments of SjC1qBP mesh
It is subcloned in carrier pET-32a (+), PCR, the prokaryotic expression plasmid of enzyme action identification restructuring occur in that in the same size with expection
DNA fragmentation (Fig. 1, right figure).Fig. 1 left figures are pcr amplification product electrophoretogram, wherein, M：DNA molecular amount mark
It is accurate；A：SjC1qBP PCR primers.Fig. 1 right figures are that double digestion identifies recombiant plasmid pET-32a (+)-SjC1qBP result figures,
Wherein, M：DNA molecular amount standard；B：The EcoR I of recombinant expression plasmid, XhoI double digestion results.To positive colony
After bacterium colony gobbet culture, bacterium solution sample is served into the sequencing of Hai Huajin biotech firms, as a result confirms nucleotide sequence coded reading
Frame is correct.
Expression of 2.2pET32-a (+)-SjC1qBP in escherichia coli and the purification of recombiant protein
Recombined pronucleus expression plasmid pET-32a (+)-SjC1qBP succeed in e. coli bl21 expression.In 1mM IPTG
After 5 hours of induction, protein expression reaches maximum (see Fig. 2, in fig. 2, M：Albumen Marker；0～8：Restructuring
Expression plasmid pET-32a (+)-SjC1qBP in e. coli bl21, at 37 DEG C, under the conditions of 1mM IPTG, induction 0,1,
2nd, protein expression situation after 3,4,5,6,7,8 hours).Recombiant protein Jing SDS-PAGE electrophoresis results after purification show
Show, purer recombiant protein has been obtained (see Fig. 3, in figure 3, M：Albumen Marker；1：Binding eluents；2-5：
Washing eluents；6-9：Elution eluents).
The detection of antigenicity of 2 Schistosoma japonicum recombiant protein rSjC1qBP of embodiment
1.Western Blotting analyze recombinant protein antigen
Recombiant protein rSjC1qBP after Ni column purifications is carried out into SDS-PAGE electrophoresis, subsequently under the conditions of 4 DEG C, is adopted
130mA, 75min are transferred on NC films, and the antigenicity of recombiant protein is analyzed as first antibody with the antiserum of anti-recombiant protein.
2.Western Blot analyze the result of recombinant protein antigen
Western Blot analysis results show that recombiant protein rSjC1qBP can be distinguished by recombiant protein immune serum and positive serum
Identification, illustrates that recombiant protein rSjC1qBP has good immunogenicity (Fig. 4).In the diagram, M：Marker；A：
One resists for mice positive serum；B：One resists for Normal Mouse Serum.
The immunization experiment of 3 Schistosoma japonicum recombiant protein rSjC1qBP (abbreviation rSjC1BP) of embodiment
1. method and step
1.1 animal immunes are tested
By 6 week old BALB/c mouse, it is divided into four groups, i.e. recombiant protein rSjC1qBP immune group, 206 vehicle control groups,
PET32-a (+) label protein group and PBS control group, per group of 10 mices.When recombiant protein immune group mice is immune every time, often
The emulsion of 100 μ L recombiant protein rSjC1qBP (20 μ g) of mouse subcutaneous injection and 206 adjuvants.206 vehicle control groups are every
The emulsion of each 100 μ L206 adjuvants of subcutaneous injection of mice and PBS.The each subcutaneous injection of every mice of PBS control group
100μLPBS.Immunity three times altogether, every minor tick 2 weeks, 7 days after immunity every time, to every mouse orbit blood sampling, are collected
Serum, saves backup in -20 DEG C.
The detection of 1.2 specific antibody level
Each immune group mice 1 week before immunity, is taken a blood sample every time, operates according to indirect ELISA method respectively for 2 weeks after immunity, with
Overnight, next day is with 1.5% bovine serum albumin (PBST dilutions) in 37 DEG C for 4 DEG C of coatings of l0 μ g/ml rSjC1qBP purification of recombinant proteins
Closing 1h, mice serum make 1：100 dilutions, incubate 1h in 37 DEG C, with HRP- goat anti-mouse iggs (1:2500) do two to resist,
1h is incubated in 37 DEG C, is washed with PBST 3 times between each step, each 5min, is finally developed the color with soluble T MB solution,
2mol/L sulphuric acid terminating reactions, read OD at 450nm wavelength450Value, detects respectively and is induced by recombiant protein rSjC1qBP
The level of specific antibody IgG of generation.
2. result
The detection of 2.1 specific antibody level
Using ELISA method detection recombinant immune group, 206 vehicle control groups, pET32-a (+) label protein group and PBS control
The change of anti-rSjC1BP specific IgG levels in group mice serum, as a result as shown in Figure 5.As can be seen from Fig. 5, for the first time
After immune recombiant protein rSjC1BP, in the mice serum of immune group, Specific IgG antibody titre is raised to higher level, and this
Kind of high level is continued until that infection cercaria is cutd open after 42 days when killing.And in the little of 206 vehicle control groups and PBS blank control groups
In Mus serum, the IgG antibody titre of the anti-rSjC1BP of specificity maintains reduced levels in whole process always, and change is not clear
It is aobvious.This explanation, immune recombiant protein can be in the specific IgG antibodies (P of induced higher levels in mice body<0.01).
Specific IgG antibodies level in 1 serum of table
4 Schistosoma japonicum recombiant protein rSjC1BP Biological Functional Assays of embodiment
1. method and step
1.1 sheep red blood cell (SRBC) hemolysis inhibition tests
1.. it is dissolved in 50 microlitres of gelatin barbitol buffer solution (SGVB2+) in same concentrations (0.01 μM～10 μM)
RSjC1BP, pET-32a label protein and BSA, with 100 microlitres of normal guinea pig serum, 37 DEG C are incubated 30 minutes.
2.. add 50 microlitres of antibody sensitizeds sheep red blood cell (SRBC) (EA, 2 × 108Individual/mL), it is uniform mix after at 37 DEG C water
Bath 30 minutes is simultaneously centrifuged (6000rpm 1min).
3.. result of the test is each group degree of hemolysis (hemoglobin of release) by determining light absorption value of the supernatant at 450nm
To determine.
4.. using bovine serum albumin as control.And while three groups of experimental system matched group is set, one group is normal haemolysis group：The
One 50 microlitres of step gelatin barbitol buffer solution (SGVB2+) in be not added with any albumen, other conditions are identical with experimental group；Two groups for not
Haemolysis group：Replace normal guinea pig serum with gelatin barbitol buffer solution, other conditions are identical with experimental group；Three groups is 100% molten
Blood group：50 microlitres EA and 150 microlitre of aquesterilisa.
1.2 Schistosoma japonicum recombiant protein rSjC1BP and C1Q. binding tests
RSjC1BP (10 μ g/ holes) is coated with overnight for 4 DEG C in ELISA Plate with 0.1M carbonate buffer solutions.Use PBST board-washings
After three times, 1h is closed in 1% gelatin of PBST dilutions at 37 DEG C.Then the C1q complements of purification are pressed into different quality dilution
In PBST, concentration is respectively 5,4,3,2,1,0.5,0.25,0 μ g/mL, closes 1h at 37 DEG C.Add after board-washing
1:The C1q complement antibodies of 2000PBST dilutions, 37 DEG C of incubation 1h.Developed the color with soluble T MB substrates, used microplate reader
Absorbance is read at 450nm wavelength.(control in this test, to be coated with the BSA of rSjC1BP same concentrations as right
According to)
2. result
2.1 hemolytic test
As a result show, bovine serum albumin can not suppress complement hemolysis, and recombiant protein rSjC1BP can suppress complement hemolysis,
The haemolysis suppression ratio of generation up to more than 30% (Fig. 6), the concentration of its suppression ratio and recombiant protein rSjC1BP within the specific limits
Into positive correlation.
2.2 complement fixation test
The combination of recombiant protein rSjC1BP and C1Q. be have detected using similar indirect ELISA method, as a result show egg of recombinating
The combination (Fig. 7) of white rSjC1BP and human complement C1q.
Embodiment described above only expresses embodiments of the present invention, and its description is more concrete and in detail, but can not therefore and
It is interpreted as the restriction to the scope of the claims of the present invention.It should be pointed out that for the person of ordinary skill of the art, do not taking off
On the premise of present inventive concept, some deformations and improvement can also be made, these belong to protection scope of the present invention.Therefore,
The protection domain of patent of the present invention should be defined by claims.
Claims (10)
1. a kind of recombinant vector, it is characterised in that comprising Schistosoma japonicum C1qBP genes, the gene order are coding SEQ ID
The nucleotide sequence of aminoacid sequence shown in NO.1.
2. recombinant vector according to claim 1, it is characterised in that the gene order is core shown in SEQ ID NO.2
Nucleotide sequence.
3. a kind of host cell comprising recombinant vector described in claim 1 or 2.
4. a kind of Schistosoma japonicum recombiant protein, it is characterised in that the recombiant protein is by recombinant vector Jing described in claim 1
Express and be obtained.
5. a kind of preparation method of Schistosoma japonicum recombiant protein, it is characterised in that comprise the following steps：
Build the recombinant expression carrier of the genes of C1qBP containing Schistosoma japonicum；
The recombinant expression carrier is transformed into into e. coli host cell；
E. coli host cell of the culture comprising the recombinant expression carrier, and under suitable conditions, induce the recombinant expressed load
Body surface Dagri japonicum C1qBP albumen.
6. preparation method according to claim 5, it is characterised in that also including step：The Japan that abduction delivering is obtained
Schistosomicide C1qBP albumen is further purified.
7. a kind of complement inhibitor, it is characterised in that comprising the Schistosoma japonicum recombiant protein described in claim 4.
8. it is a kind of can with described in claim 4 Schistosoma japonicum recombiant protein specific binding antibody.
9. application of the Schistosoma japonicum recombiant protein described in claim 4 in treatment schistosomicide medicine is prepared.
10. application of the Schistosoma japonicum recombiant protein described in claim 4 in the medicine for preparing treatment complement abnormality disease.
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