CN109810959A - It is a kind of in conjunction with KEAP1 and to adjust the polypeptide of NRF2 protein stability - Google Patents
It is a kind of in conjunction with KEAP1 and to adjust the polypeptide of NRF2 protein stability Download PDFInfo
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Abstract
It is specifically a kind of in conjunction with KEAP1 and to adjust the polypeptide of NRF2 protein stability the present invention relates to field of biomedicine.Oxidative stress regulation plays a significant role during tumor development, as cellular anti-oxidant key molecule, the factor for influencing NRF2 expression and protein stability all plays a significant role NRF2 for adjusting cellular oxidation stress tolerance ability and chemotherapeutics repellence.The present invention proposes the polypeptide of competitive binding KEAP1 a kind of on the basis of RMP (RNA polymerase II subunit 5-mediating protein), it improves cellular anti-oxidant capacity by enhancing NRF2 protein stability.The present invention provides useful targeting target to regulation cellular anti-oxidant capacity.
Description
Technical field
The present invention relates to fields of biomedicine, specifically, being one kind in conjunction with KEAP1 and adjusting NRF2 protein stability
Polypeptide, polypeptide competitive binding KEAP1, make cellular anti-oxidant regulation key protein NRF2 ubiquitination degradation subtract
It is few, to enhance tumour oxidation resistance.
Background technique
RMP (RNA polymerase II subunit 5-mediating protein, RMP are referred to as URI,
Unconventional prefoldin RPB5 Interactor) it is to find HBx by Japanese researchers in 1998 to turn earliest
Record is found during adjusting GAP-associated protein GAP: the 5th subunit of RNA polymerase II (RNA polymerase II subunit 5,
RPB5 it) is bound directly with HBx, while the two passes through respective binding site in conjunction with II B of TF, the complex that this three is formed promotees
Into the transcription of HBx；And they have found RMP in the lower situation of HBx content, competitive binding RBP5 covers the latter's protein structure
In with II B binding site of HBx and TF, to inhibit the transcription of HBx.2011, Switzerland scientist reported that URI exists as oncogene
Tumor acceleration is played in oophoroma: URI forms complex in conjunction with PP1 γ and inhibits its phosphatase activity, to destroy
The negative feedback path of the latter and S6K1-BAD composition, BAD phosphorylation increase, and Apoptosis is reduced, and survival increases
(J.P.Theurillat, S.C.Metzler, N.Henzi, et al.URI is an oncogene amplified in
ovarian cancer cells and is required for their survival[J].Cancer Cell,2011,
19(3):317-32.).Then, more research discovery RMP are all kinds of in prostate cancer, Huppert's disease, carcinoma of endometrium etc.
Tumor acceleration is played in malignant tumour.In addition, also having been reported that discovery RMP can easily change p65 in conjunction with IL-6 promoter region, promote
It is transcribed, and the more IL-6 of secretion of hepatoma are made, and by IL-6/STAT3 signal path, promotes hepatoma Metastasis and self-renewing.
ROS (reactive oxygen species) plays a significant role tumor development.It is intracellular excessive
ROS causes key gene mutation that cell Proliferation and tumour is promoted to occur.Tumour during the growth process, needs to cope with various inner and outer rings
The variation of border factor.Wherein ROS is largely generated and is discharged, and cellular redox equilibrium state can be caused to break, and is caused a series of
Phenomena such as such as DNA damage, membrane structure destroy, organelle destroys, eventually leads to the generation of Apoptosis.Therefore, it explores
Cellular anti-oxidant regulatory mechanism during tumor development, has very important significance.
KEAP1-NRF2-ARE signal path is played an important role during cellular anti-oxidant.KEAP1(Kelch-like
ECH-associated protein 1) it is anchored in cytoplasm with actin, while the region Kelch and NRF2 can be passed through
In conjunction with.KEAP1 provides bracket for E3 ubiquitin ligase CUL3, and the latter can mediate NRF2 degradation by Ubiquitin-proteasome.
Under normal circumstances, by above-mentioned mechanism, NRF2 only maintains reduced levels, to guarantee daily vital movement.When oxidative stress, upstream
Related stimulus mediates NRF2 degradation to terminate and accumulate, and eventually enters into nucleus.Enter the NRF2 and Antioxidant responsive element of core
(antioxidant response element (ARE)) is combined, and promotes the transcription of anti-oxidant related gene, participates in cell antioxygen
Change in relevant every physiological activity.(John P.Fruehauf, Frank L.Meyskens, Jr.Reactive oxygen
Species:A breath of life or death? [J] .Clinical Cancer Research, 2007,13 (3):
789-794.)
Targetedly intervene important molecule in anti-oxidant related pathways, important work is all had for tumour occurrence and development
With also for oncotherapy offer personalised drug guidance.
Summary of the invention
It being capable of competitive binding KEAP1, stable NRF2 the purpose of the present invention is to provide one section in a kind of RMP protein molecular
Albumen improves the polypeptide RMP-D2 of cellular anti-oxidant capacity.The present invention also provides the codings of above-mentioned polypeptide RMP-D2
Gene, the recombinant vector containing above-mentioned encoding gene, recombinant bacterium, recombinant cell or expression cassette, and its tumour is being adjusted in preparation
Application in the drug of oxidation resistance.
To achieve the goals above, the first aspect of the present invention provides a kind of RMP-D2 polypeptide, and amino acid sequence is such as
(I) or shown in (II):
(I) amino acid sequence as shown in SEQ ID NO.1；
(II) amino acid sequence as shown in SEQ ID NO.1 is modified, replaces, misses or adds one or several amino
The amino acid sequence that acid obtains.
RMP albumen previously research mainly think its by forming complex in conjunction with PP1 γ and inhibiting its phosphatase activity,
To destroy the negative feedback path of the latter and S6K1-BAD composition, BAD phosphorylation increases, and Apoptosis is reduced, and survival increases.
The mechanism is used to interpret the reason of RMP promotes kinds of tumors progress.But the present inventor in long-term research process for the first time
It was found that RMP albumen passes through the region-competitive combination KEAP1 containing E**E structural domain between its protein molecular 164-288 amino acid
NRF2 binding site in protein molecular enhances NRF2 protein stability, and then regulating cell oxidation resistance.The RMP albumen point
The peptide fragment amino acid sequence and nucleotide sequence of sub- 164-288 amino acid composition are as shown in SEQ ID NO.1.It needs to illustrate
It is by the substitution of one or several amino acid residues and/or missing and/or to add amino acid sequence shown in SEQ ID NO.1
Add, and the derived sequence with cellular anti-oxidant capacity, also belongs to protection scope of the present invention.
The second aspect of the present invention provides a kind of encoding gene of RMP-D2 polypeptide as described above, nucleotide sequence
As shown in (i) or (ii):
(i) nucleotide sequence as shown in SEQ ID NO.2；
(ii) nucleotide sequence as shown in SEQ ID NO.2 is modified, replaces, misses or adds one or more bases
The nucleotide sequence of acquisition.
It displacement and/or missing and/or insertion by above-mentioned nucleotide sequence Jing Guo one or several bases and encodes out
Peptide fragment has the derived sequence of the identical oxidation resistance with the peptide fragment of above-mentioned amino acid sequence, also belongs to protection model of the invention
It encloses.
The third aspect of the present invention provides a kind of recombinant vector, recombinant bacterium, recombinant cell or expression cassette, contains as above
The encoding gene of the RMP-D2 polypeptide.
Further, the recombinant vector is one in slow virus carrier, retroviral vector or adenovirus vector
Kind.
Further, the recombinant vector contains described in promotion in the upstream of the encoding gene of the polypeptide or downstream
Encoding gene expression enhancer (being often referred to composing type specific enhancers such as CMV, SV40, PGK enhancer etc.).
The fourth aspect of the present invention provides a kind of RMP-D2 polypeptide as described above and is preparing regulating cell oxidation resistance
Application in drug.
Further, the RMP-D2 polypeptide, can competitive binding KEAP1, thus reduce NRF2 degradation, enhancing
NRF2 protein stability improves cellular anti-oxidant capacity.
The present invention also provides a kind of RMP-D2 polypeptides as described above to prepare competitive binding KEAP1, reduces NRF2 drop
Solution enhances NRF2 protein stability, improves the application in cellular anti-oxidant capacity drug.
The fifth aspect of the present invention, the encoding gene for providing a kind of RMP-D2 polypeptide as described above are preparing regulating cell
Application in oxidation resistance drug.
The sixth aspect of the present invention provides a kind of recombinant vector as described above, recombinant bacterium, recombinant cell or expression cassette and exists
Prepare the application in regulating cell oxidation resistance drug.
The seventh aspect of the present invention provides a kind of RMP-D2 polypeptide as described above, encoding gene, recombinant vector, recombination
Bacterium, recombinant cell or expression cassette application in preparation of anti-tumor drugs.
The eighth aspect of the present invention provides a kind of RMP-D2 polypeptide as described above, encoding gene, recombinant vector, recombination
Bacterium, recombinant cell or expression cassette assist alleviating the application in drug of the tumour cell to other chemotherapeutics drug resistances in preparation.
Further, the tumour is to be primary in the various types tumour of liver, especially various types cholangiocarcinoma.
The ninth aspect of the present invention, provides a kind of drug of regulating cell oxidation resistance, and the drug includes as above
RMP-D2 polypeptide, encoding gene, recombinant vector, recombinant bacterium, recombinant cell or the expression cassette.Further, the medicine
Object also includes pharmaceutically acceptable auxiliary material.
The tenth aspect of the present invention, provides a kind of anti-tumor drug, and the drug includes RMP-D2 as described above more
Peptide, encoding gene, recombinant vector, recombinant bacterium, recombinant cell or expression cassette.Further, the drug also includes pharmaceutically
Acceptable auxiliary material.
The invention has the advantages that:
Polypeptide provided by the invention, that is, RMP-D2 polypeptide, can competitive binding KEAP1, thus reduce NRF2 degradation, increase
Strong NRF2 protein stability, improves cellular anti-oxidant capacity.It can be used as the target spot of regulating cell oxidation resistance, can apply
In prepare anti-tumor drug or auxiliary alleviate tumour cell to other chemotherapeutics drug resistances.
Oxidative stress regulation plays a significant role during tumor development, and NRF2 is as cellular anti-oxidant key point
Son, the factor for influencing NRF2 expression and protein stability are resisted for adjusting cellular oxidation stress tolerance ability and chemotherapeutics
Property all plays a significant role.The present invention proposes the polypeptide of competitive binding KEAP1 a kind of on the basis of RMP, it passes through increasing
Strong NRF2 protein stability, improves cellular anti-oxidant capacity.The present invention provides useful target to regulation cellular anti-oxidant capacity
To target.
Detailed description of the invention
Fig. 1 is that the KEAP1 albumen of the embodiment of the present invention can respectively and the combination of RMP albumen and NRF2 albumen, but RMP
There is no interaction relationship with NRF2 albumen.
Fig. 2 is that the RMP albumen of the embodiment of the present invention passes through its region D2 in conjunction with KEAP1.
Fig. 3 is that 2 region of different plant species RMP protein D of the embodiment of the present invention is similar to NRF2 protein molecular in the presence of conservative
Middle KEAP1 binding site E**E.
Fig. 4 is that body can be truncated with the KEAP1 containing Kelch repetitive structure domain in the RMP protein molecular of the embodiment of the present invention
Plasmid be combined with each other.
Fig. 5 is that the RMP protein molecular of the embodiment of the present invention can be with NRF2 protein molecular competitive binding KEAP1 albumen.
Specific embodiment
It in order to make the object, technical scheme and advantages of the embodiment of the invention clearer, below will be in the embodiment of the present invention
Technical solution carry out clear, complete description.The person that is not specified actual conditions in embodiment, according to normal conditions or manufacturer
It is recommended that condition carry out.
Cholangiocarcinoma cell system HuCCT1 and HEK-293T cell purchase cell institute, the Chinese Academy of Sciences.PCDNA3.1A-GFP is wild
Type pCDNA3.1A-RMP plasmid and corresponding truncate plasmid are purchased from Shanghai Duanxi inkstone biotech company.KEAP1 truncates constitution
Grain is by New Jersey's medicament and dentist university (The University of Medicine and Dentistry of New
Jersey, UMDNJ) professor Xia Bin provide (can also be prepared according to following documents method: J.Ma, H.Cai, T.Wu, et
al.PALB2 interacts with KEAP1 to promote NRF2 nuclear accumulation and
function[J].Mol Cell Biol,2012,32(8):1506-1).Factory is not specified in other agents useful for same or instrument
Shang Zhe is the conventional products that can be obtained by commercially available purchase.
Embodiment 1:KEAP1 albumen can respectively and the combination of RMP albumen and NRF2 albumen, but RMP and NRF2 albumen
There is no interaction relationship.
It takes out Protein A/G agarose magnetic bead (MAg25K/Protein A/G, Ying Ruicheng biochemical technology, P28-002)
First carry out magnetic bead pretreatment according to its specification: pipettor piping and druming or vortex oscillator mix magnetic bead, take 50 μ L bead suspensions
(10%, v/v) into centrifuge tube, the removal of 1.5mL magnetic frame Magnetic Isolation saves liquid；It is (reverse that 100 μ L PBS mixing magnetic bead is added
30 seconds or the mixing of vortex oscillator), Magnetic Isolation removes supernatant (being repeated 2 times)；Antibody combination is carried out again: with 100 μ
LPBST (PBST, pH 7.4:PBS with 0.02%Tween-20) dilutes 2~20 μ g antibody samples, and dress is added to after dilution
Have in the centrifuge tube of magnetic bead；By said mixture be put into shaking table or rotary mixer room temperature or 37 DEG C be mixed by inversion or be vortexed it is mixed
Even 10~15 minutes；Centrifuge tube equipped with magnetic bead and antibody is put into magnetic frame, after magnetic bead is completely adherent, removes supernatant；
It is washed magnetic bead-antibody complex 3 times with 100 μ LPBST.Antigen precipitates reaction: magnetic bead-antibody Antigen adsorption: is housed one step up
100~1000 μ L of cell pyrolysis liquid prepared is added in the centrifuge tube of compound, is blown and beaten and is mixed with liquid-transfering gun；37 DEG C reverse mixed
Even or low speed vortex mixes 15~20 minutes, combines the magnetic bead of antigen and binding antibody；By the centrifuge tube of previous step and mix
It closes object and is put into magnetic frame, draw supernatant (discardable also can be reserved for of supernatant does subsequent analysis)；Take 200 μ L PBS washing magnetic bead-anti-
Body-antigenic compound 3 times；Magnetic bead is resuspended with 100 μ L PBS after the completion of washing, for operating in next step.Antigen elution: it will be equipped with
Magnetic bead-Antibody-antigen complex centrifuge tube is put into magnetic frame, sucks supernatant after magnetic bead is adherent completely；To above-mentioned centrifuge tube
Middle addition 25 μ L Elution Buffer (100mM Glycine, pH 2.8) and 5 μ L 5x SDS-PAGE Loading
Compound is resuspended with pipettor again in Buffer；95~100 DEG C are boiled 5~10 minutes；Centrifuge tube after boiling is put into magnetic frame
Supernatant is taken to do subsequent analysis.
Collection is incubated at 10cm culture dish HuCCT1 cell, and 1ml cell pyrolysis liquid, BCA egg is added after physiological saline cleaning
It is white it is quantitative after, cell pyrolysis liquid is divided into two parts of protein content 1600ug, appropriate protein lysate is added and is settled to volume
For 500ul, residual protein makes Input group.According to above-mentioned co-immunoprecipitation experiment step, respectively using Rabbit IgG and
KEAP1 antibody is incubated for.Product is tested by Western Blotting after immunoprecipitation, will using SDS-PAGE separation gel
Different molecular weight albumen is separated, transferring film to NC (nitrocellulose) film, primary antibody and secondary antibody be incubated for after Odyssey CLx at
As system (LICOR) is analyzed.KEAP1 antibody shown in Figure 1A can be by NRF2 but also by RMP protein molecular from cholangiocarcinoma
Co-immunoprecipitation in HuCCT1 cell protein lysate.Figure 1B is by KEAP1 protein molecular using RMP antibody from cholangiocarcinoma
Co-immunoprecipitation in HuCCT1 cell protein lysate.Fig. 1 C is that HuCCT1 extracellularly turns on KEAP1 plasmid basic while transfecting
PcDNA3.1A GFP, RMP plasmid, gives 250 μM of H after 36 hours2O2Processing 8 hours, turns RMP cell outside and is able to observe that
KEAP1 increases in conjunction with RMP, and reduces in conjunction with NRF2.
Embodiment 2:RMP albumen is by its region D2 in conjunction with KEAP1.
According to each functional areas function of the past document report RMP albumen, it is designed to the truncate containing different structure territory
(D.Dorjsuren, Y.Lin, W.Wei, et al.RMP, a novel RNA polymerase II subunit5-
interacting protein,counteracts transactivation by hepatitis B virus X
protein[J].Mol Cell Biol,1998,18(12):7546-55；J.P.Theurillat, S.C.Metzler,
N.Henzi, et al.URI is an oncogene amplified in ovarian cancer cells and is
required for their survival[J].Cancer Cell,2011,19(3):317-32.；Stefan Bur é n,
Ana L.Gomes, Ana Teijeiro, et al.Regulation of OGT by URI in Response to
Glucose Confers c-MYC-Dependent Survival Mechanisms[J].Cancer Cell,2016,30
(2):290-307.).RMP truncate shown in Fig. 2A and mutant plasmid schematic diagram.It is prepared by above-mentioned truncate plasmid
Corresponding cDNA segment is analyzed according to protein fragments, designs effective primer, using HindIII-ApaI as cloning site,
PcDNA3.1 (+)/myc-His is connect as carrier, by carrier with segment, and selection is effectively cloned and expanded.Identify above-mentioned plasmid
After expression effect, using PEI transfection reagent by above-mentioned each truncate plasmid, pcDNA3.1 (+)/myc-His GFP and RMP plasmid
It is transfected together with wild KEAP1 plasmid respectively to 293T cell.After culture 36 hours, albumen is collected, it is quantitative, according to embodiment 1
The step carries out co-immunoprecipitation experiment.Wild type KEAP1 plasmid shown in Fig. 2 B can only be truncated with the RMP containing the region D2
The interaction of body (D2, D4, D5) protein molecular.
Embodiment 3: there is the conservative KEAP1 in NRF2 protein molecular that is similar to and combine in 2 region of different plant species RMP protein D
Site E**E.
Lot of documents is reported in the cohesive process of KEAP1 and NRF2, and it is (strong to combine to be present in ETGE in NRF2Neh2 structural domain
Site) and DLG (weak binding site) be important KEAP1 binding site (M.Rojo de la Vega, E.Chapman,
D.D.Zhang.NRF2and the Hallmarks of Cancer[J].Cancer Cell,2018,34(1):21-43；
Hanna M.Leinonen, Emilia Kansanen, Petriet al.Role of the Keap1–
Nrf2Pathway in Cancer[J].2014,122:281-320；P.Canning, F.J.Sorrell,
A.N.Bullock.Structural basis of Keap1interactions with Nrf2[J].Free Radic
Biol Med,2015,88(Pt B):101-107.).By NCBI inquirer's RMP protein sequence, only there are similar in the region D2
In ETGE sequence.Inquire mouse, Norway Rat, zebra fish, orangutan, RMP protein sequence in the species such as chicken again, by above-mentioned sequence into
Row compares analysis, and the region D2 of these as shown in Figure 3 species is relatively conservative region, and the region D2 in these protein sequences
Two containing similar to KEAP1 binding site E**E in NRF2 molecule.The amino acid sequence of RMP albumen such as SEQ ID in Fig. 3
Shown in NO.3-SEQ ID NO.8.
Embodiment 4:RMP protein molecular can be with NRF2 protein molecular competitive binding KEAP1 albumen.
HEK-293T extracellularly turns NRF2, KEAP1 plasmid while gradient transfected wild-type RMP plasmid (2 μ g, 6 μ g, 12 μ
G), after transfecting 36 hours, albumen is collected, the carry out co-immunoprecipitation passes through as shown in Fig. 4 A, 4B in accordance with the above-mentioned embodiment 1
Analysis finds that as RMP expression quantity increases, competitiveness increases in conjunction with KEAP1, and NRF2 is combined and reduced.
Embodiment 5:RMP protein molecular can influence NRF2 protein stability.
It is overexpressed HuCCT1 cell line by slow virus system construction RMP, uses protein synthesis inhibitor CHX
(Cycloheximide) control and overexpressing cell are handled according to time gradient 0,15,30,60,120,180min.Albumen is added to split
It solves liquid and collects albumen, BCA is quantitative, prepares loading sample, is sent out as shown in Fig. 5 A, 5B by Western Blotting experimental analysis
Existing RMP, which is overexpressed, inhibits NRF2 protein degradation.
The preferred embodiment of the present invention has been described in detail above, but the invention be not limited to it is described
Embodiment, those skilled in the art can also make various equivalent on the premise of not violating the inventive spirit of the present invention
Variation or replacement, these equivalent variation or replacement are all included in the scope defined by the claims of the present application.
Sequence table
<110>Second Military Medical University, PLA
<120>a kind of in conjunction with KEAP1 and to adjust the polypeptide of NRF2 protein stability
<130> /
<160> 8
<170> SIPOSequenceListing 1.0
<210> 1
<211> 125
<212> PRT
<213>artificial sequence (Artificial)
<400> 1
Glu Ile Lys Cys Asp Phe Glu Phe Lys Ala Lys His Arg Ile Ala His
1 5 10 15
Lys Pro His Ser Lys Pro Lys Thr Ser Asp Ile Phe Glu Ala Asp Ile
20 25 30
Ala Asn Asp Val Lys Ser Lys Asp Leu Leu Ala Asp Lys Glu Leu Trp
35 40 45
Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Leu Leu Gly Glu Leu
50 55 60
Asp Ser Lys Pro Asp Thr Val Ile Ala Asn Gly Glu Asp Thr Thr Ser
65 70 75 80
Ser Glu Glu Glu Lys Glu Asp Arg Asn Thr Asn Val Asn Ala Met His
85 90 95
Gln Val Thr Asp Ser His Thr Pro Cys His Lys Asp Val Ala Ser Ser
100 105 110
Glu Pro Phe Ser Gly Gln Val Asn Ser Gln Leu Asn Cys
115 120 125
<210> 2
<211> 375
<212> DNA
<213>artificial sequence (Artificial)
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gaaattaaat gtgacttcga atttaaagca aaacaccgaa ttgctcataa accgcattcc 60
aaaccaaaaa cttcagatat ttttgaagca gatattgcaa atgatgtgaa atccaaggat 120
ttgctagctg ataaagaact gtgggctcga cttgaagaac tagagagaca ggaagaattg 180
ctgggtgaac ttgatagtaa gcctgatact gtgattgcaa atggagaaga tacgacatct 240
tctgaagagg aaaaggaaga tcgtaacaca aatgtgaatg cgatgcatca agtaacagac 300
tctcatactc cttgtcataa ggatgttgca agttcagaac cattcagtgg tcaagtgaat 360
agtcagttga actgt 375
<210> 3
<211> 53
<212> PRT
<213>homo sapiens (Homo sapiens)
<400> 3
Glu Leu Trp Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Leu Leu
1 5 10 15
Gly Glu Leu Asp Ser Lys Pro Asp Thr Val Ile Ala Asn Gly Glu Asp
20 25 30
Thr Thr Ser Ser Glu Glu Glu Lys Glu Asp Arg Asn Thr Asn Val Asn
35 40 45
Ala Met His Gln Val
50
<210> 4
<211> 51
<212> PRT
<213>mouse (Mus musculus)
<400> 4
Glu Leu Trp Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Leu Leu
1 5 10 15
Gly Glu Leu Glu Ser Lys Pro Asp Thr Val Ile Ala Asn Gly Glu Asp
20 25 30
Arg Val Ser Ser Glu Glu Glu Lys Glu Gly Ala Asp Thr Gly Val Asn
35 40 45
Val Val Ser
50
<210> 5
<211> 52
<212> PRT
<213>Norway Rat (Rattus norvegicus)
<400> 5
Glu Leu Trp Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Leu Leu
1 5 10 15
Gly Glu Leu Lys Ser Lys Pro Asp Thr Val Ile Ala Asn Gly Glu Asp
20 25 30
Thr Val Ser Ser Glu Glu Glu Lys Glu Asp Glu Asp Thr Gly Val Asn
35 40 45
Val Val Ser Ser
50
<210> 6
<211> 52
<212> PRT
<213>zebra fish (Danio rerio)
<400> 6
Glu Leu Trp Ala Arg Leu Asp Glu Leu Glu Arg Gln Glu Glu Leu Gln
1 5 10 15
Asp Gln Arg Phe Arg Leu Asp Ser Thr Asp Thr Asn Gly Glu Asp Thr
20 25 30
Thr Ser Ser Ser Glu Glu Glu Lys Glu Ala Asp Gly Gly Ser Asp Val
35 40 45
Gln Val Asn His
50
<210> 7
<211> 52
<212> PRT
<213>chimpanzee (Pan troglodytes)
<400> 7
Glu Leu Trp Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Leu Leu
1 5 10 15
Gly Glu Leu Asp Ser Lys Pro Asp Thr Val Ile Ala Asn Gly Glu Asp
20 25 30
Thr Thr Ser Ser Glu Glu Glu Lys Glu Asp Arg Asn Thr Asn Val Asn
35 40 45
Ala Met His Gln
50
<210> 8
<211> 52
<212> PRT
<213>chicken (Gallus gallus)
<400> 8
Glu Leu Trp Ala Arg Leu Glu Glu Leu Glu Arg Gln Glu Glu Ile Leu
1 5 10 15
Gly Glu Leu Asp Arg Met Pro Asp Thr Val Glu Thr Asn Gly Glu Asp
20 25 30
Thr Thr Ser Ser Glu Glu Glu Lys Glu Asp Lys Arg Met Asp Leu Asn
35 40 45
Gly Thr Tyr Arg
50
Claims (10)
1. a kind of RMP-D2 polypeptide, amino acid sequence is such as shown in (I) or (II):
(I) amino acid sequence as shown in SEQ ID NO.1；
(II) amino acid sequence as shown in SEQ ID NO.1 is modified, replaces, misses or adds one or several amino acid and obtain
The amino acid sequence obtained.
2. a kind of encoding gene of RMP-D2 polypeptide as described in claim 1, nucleotide sequence is such as shown in (i) or (ii):
(i) nucleotide sequence as shown in SEQ ID NO.2；
(ii) nucleotide sequence as shown in SEQ ID NO.2 is modified, replaces, misses or adds one or more bases acquisitions
Nucleotide sequence.
3. a kind of recombinant vector, recombinant bacterium, recombinant cell or expression cassette contain RMP-D2 polypeptide as claimed in claim 2
Encoding gene.
4. recombinant vector according to claim 3, recombinant bacterium, recombinant cell or expression cassette, which is characterized in that the weight
Group carrier is one of slow virus carrier, retroviral vector or adenovirus vector.
5. recombinant vector according to claim 4, recombinant bacterium, recombinant cell or expression cassette, which is characterized in that the weight
Group carrier contains the enhancer for promoting the encoding gene expression in the upstream of the encoding gene of the polypeptide or downstream.
6. RMP-D2 polypeptide as described in claim 1, the encoding gene of RMP-D2 polypeptide as claimed in claim 2 are such as weighed
Benefit require 3 described in recombinant vector, recombinant bacterium, recombinant cell or expression cassette in preparing regulating cell oxidation resistance drug
Using.
7. RMP-D2 polypeptide as described in claim 1, the encoding gene of RMP-D2 polypeptide as claimed in claim 2 are such as weighed
Benefit require 3 described in recombinant vector, recombinant bacterium, recombinant cell or expression cassette application in preparation of anti-tumor drugs.
8. RMP-D2 polypeptide as described in claim 1, the encoding gene of RMP-D2 polypeptide as claimed in claim 2 are such as weighed
Benefit require 3 described in recombinant vector, recombinant bacterium, recombinant cell or expression cassette preparation assist alleviate tumour cell to other chemotherapy
Application in the drug of drug resistance.
9. a kind of drug of regulating cell oxidation resistance, the drug includes RMP-D2 polypeptide as described in claim 1,
The encoding gene of RMP-D2 polypeptide as claimed in claim 2, recombinant vector as claimed in claim 3, recombinant bacterium, recombination are thin
Born of the same parents or expression cassette.
10. a kind of anti-tumor drug, the drug includes RMP-D2 polypeptide as described in claim 1, such as claim 2 institute
The encoding gene for the RMP-D2 polypeptide stated, recombinant vector, recombinant bacterium, recombinant cell or expression cassette as claimed in claim 3.
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Cited By (2)
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| CN111358936A (en) * | 2020-03-20 | 2020-07-03 | 中国人民解放军63919部队 | Application of NRF2 protein in preparing medicine for regulating biological rhythm |
| CN112210561A (en) * | 2020-11-06 | 2021-01-12 | 浙江大学 | Application of OsKEAP1 gene in regulation of rice seed phenotype and germination rate |
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| WO2001085762A2 (en) * | 2000-05-12 | 2001-11-15 | Novartis Forschungsstiftung Zweigniederlassung Friedrich Miescher Institute For Biomedical Research | Cancer diagnosis and assays for screening anti-cancer agents |
| TWI629986B (en) * | 2017-06-27 | 2018-07-21 | 台灣利得生物科技股份有限公司 | Use of ricotinic acid M for preparing a pharmaceutical composition having anti-aging activity against high sugar |
| CN110787180A (en) * | 2019-12-19 | 2020-02-14 | 济南大学 | Application of homoplantaginoside and derivatives thereof as Nrf-2 activator |
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2019
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| WO2001085762A2 (en) * | 2000-05-12 | 2001-11-15 | Novartis Forschungsstiftung Zweigniederlassung Friedrich Miescher Institute For Biomedical Research | Cancer diagnosis and assays for screening anti-cancer agents |
| TWI629986B (en) * | 2017-06-27 | 2018-07-21 | 台灣利得生物科技股份有限公司 | Use of ricotinic acid M for preparing a pharmaceutical composition having anti-aging activity against high sugar |
| CN110787180A (en) * | 2019-12-19 | 2020-02-14 | 济南大学 | Application of homoplantaginoside and derivatives thereof as Nrf-2 activator |
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| DORJBAL DORJSUREN ET AL.: "RMP,a novel RNA polymerase II subunit 5-interacting protein,counteracts transactivation by hepatitis B virus X protein", 《MOLECULAR AND CELLULAR BIOLOG》 * |
| GENBANK: "PREDICTED: Homo sapiens URI1, prefoldin like chaperone (URI1), transcript variant X2,mRNA NCBI Reference Sequence: XM_005259363.4", 《GENBANK》 * |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN111358936A (en) * | 2020-03-20 | 2020-07-03 | 中国人民解放军63919部队 | Application of NRF2 protein in preparing medicine for regulating biological rhythm |
| CN112210561A (en) * | 2020-11-06 | 2021-01-12 | 浙江大学 | Application of OsKEAP1 gene in regulation of rice seed phenotype and germination rate |
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| CN109810959B (en) | 2022-03-15 |
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