CN117147866A - Application of muscle-source 6 phosphofructokinase and polypeptide fragment thereof in urine or urine exosomes in diabetes - Google Patents

Application of muscle-source 6 phosphofructokinase and polypeptide fragment thereof in urine or urine exosomes in diabetes Download PDF

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CN117147866A
CN117147866A CN202210429495.4A CN202210429495A CN117147866A CN 117147866 A CN117147866 A CN 117147866A CN 202210429495 A CN202210429495 A CN 202210429495A CN 117147866 A CN117147866 A CN 117147866A
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ala
leu
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张曼
王佶图
孟倩
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    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
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    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6893—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids related to diseases not provided for elsewhere

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Abstract

The invention provides application of muscle-source 6 phosphofructokinase (ATP-dependent 6-phosphofructokinase, muscle type) and polypeptide fragments thereof in urine or urine exosomes, in particular application of the muscle-source 6 phosphofructokinase and polypeptide fragments thereof in urine or urine exosomes in preparation of preparations for diagnosis, differential diagnosis, disease degree judgment, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like. The research proves that the expression of the muscle-source 6 phosphofructokinase and the polypeptide fragment thereof in the urine or urine exosomes is reduced in diabetics compared with healthy people (normal control group). Can be used for various purposes of diabetics. The invention plays the advantages of noninvasive acquisition of urine specimen, large-scale repeated sampling and convenient preservation, and utilizes the urine specimen to detect the muscle-source 6 phosphofructokinase and the polypeptide fragments thereof in urine or urine exosomes.

Description

Application of muscle-source 6 phosphofructokinase and polypeptide fragment thereof in urine or urine exosomes in diabetes
Technical Field
The invention relates to new application of muscle-source 6 phosphofructokinase and polypeptide fragments thereof in urine or urine exosomes, in particular to application of the muscle-source 6 phosphofructokinase and polypeptide fragments thereof in preparation of diabetes diagnosis, differential diagnosis, disease degree judgment, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like.
Background
Diabetes mellitus is a metabolic endocrine disorder characterized by a disturbance in glucose and fat metabolism and an increase in plasma glucose levels, resulting from the combined action of genetic and environmental factors. In recent years, the global population has a rapidly growing incidence of diabetes mellitus, which has become the most challenging worldwide health problem in the 21 st century. Along with the improvement of living standard and the transition of living style, the incidence rate of diabetes in China is in a rapid rising trend, and the statistical result shows that the number of diabetes patients in China is the second in the world at present, and the diabetes population in 2040 years is expected to be as high as 6.42 hundred million.
Diabetes hyperglycemia can lead to metabolic disorders of carbohydrates, proteins and fats, leading to neurological, renal and cardiovascular complications, which are a great potential for public health problems. Diabetes mellitus is hidden, and most patients cannot find the diabetes mellitus timely, so that the diabetes mellitus is usually diagnosed due to clinical manifestations of complications. In addition, diabetics have a long course of disease, need to monitor blood sugar for a long time or even for life, and repeatedly draw blood and perform invasive examination to cause a plurality of inconveniences for the patients. However, if diabetes patients are not diagnosed in time, a plurality of viscera damage occurs in the whole body along with the progress of the disease, and serious patients are disabled and die even due to complications. In summary, diabetes not only causes inconvenience and pain to patients, but also brings heavy burden to families and society of patients. In order to improve the life quality of diabetics and solve the practical problems of the diabetics, it is necessary to research a noninvasive, convenient, quick and easily repeatable diabetes inspection method.
Muscle-derived phosphofructokinase 6 (ATP-PFK) is a key enzyme in sugar metabolism, plays a vital role in the sugar metabolism of the body, and is an important step in human glycolysis. In the present study, the content of muscle-derived phosphofructokinase 6 in urine or urine exosomes of diabetics was down-regulated compared with the healthy human group, and the content was reduced.
Compared with the common clinical blood sample, the urine can be completely and noninvasively collected continuously in a large amount; without steady state regulation, more kinds and larger amplitude of changes can be accumulated, and many pathophysiological changes of the body may be manifested in urine. Protein polypeptides with relatively small molecular weights such as hormones and cytokines can be excreted into urine quickly after entering blood, and the probability of being detected in the urine is much higher than that in the blood; prior to urine collection, the possible proteolytic processing of urine is completed, so that urine proteins remain stable for a longer period of time. In order to relieve the pain of multiple blood sampling of diabetics, the experiment is expected to realize painless, convenient, quick and easily repeated urine detection to assist diagnosis and disease monitoring of the diabetics through urine protein or polypeptide research on the basis of early-stage methodology, and lays a foundation for further research of a urine polypeptide detection kit.
Disclosure of Invention
The invention aims to provide an application of muscle-source 6 phosphofructokinase and polypeptide fragments thereof in urine or urine exosomes in preparation of preparations for diagnosis of diabetes mellitus, differential diagnosis, judgment of disease degree, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like.
Preferably, the amino acid sequence of the muscle-derived phosphofructokinase 6 in the urine or urine exosomes comprises the amino acid sequence shown in SEQ ID NO.1 (MTHEEHHAAK TLGIGKAIAV LTSGGDAQGM NAAVRAVVRV GIFTGARVFF VHEGYQGLVD GGDHIKEATW ESVSMMLQLG GTVIGSARCK DFREREGRLR AAYNLVKRGI TNLCVIGGDG SLTGADTFRS EWSDLLSDLQ KAGKITDEEA TKSSYLNIVG LVGSIDNDFC GTDMTIGTDS ALHRIMEIVD AITTTAQSHQ RTFVLEVMGR HCGYLALVTS LSCGADWVFI PECPPDDDWE EHLCRRLSET RTRGSRLNII IVAEGAIDKN GKPITSEDIK NLVVKRLGYD TRVTVLGHVQ RGGTPSAFDR ILGSRMGVEA VMALLEGTPD TPACVVSLSG NQAVRLPLME CVQVTKDVTK AMDEKKFDEA LKLRGRSFMN NWEVYKLLAH VRPPVSKSGS HTVAVMNVGA PAAGMNAAVR STVRIGLIQG NRVLVVHDGF EGLAKGQIEE AGWSYVGGWT GQGGSKLGTK RTLPKKSFEQ ISANITKFNI QGLVIIGGFE AYTGGLELME GRKQFDELCI PFVVIPATVS NNVPGSDFSV GADTALNTIC TTCDRIKQSA AGTKRRVFII ETMGGYCGYL ATMAGLAAGA DAAYIFEEPF TIRDLQANVE HLVQKMKTTV KRGLVLRNEK CNENYTTDFI FNLYSEEGKG IFDSRKNVLG HMQQGGSPTP FDRNFATKMG AKAMNWMSGK IKESYRNGRI FANTPDSGCV LGMRKRALVF QPVAELKDQT DFEHRIPKEQ WWLKLRPILK ILAKYEIDLD TSDHAHLEHI TRKRSGEAAV), SEQ ID NO.2 (MTHEEHHAAK TLGIGKAIAV LTSGGDAQGM NAAVRAVVRV GIFTGARVFF VHEGYQGLVD GGDHIKEATW ESVSMMLQLG GTVIGSARCK DFREREGRLR AAYNLVKRGI TNLCVIGGDG SLTGADTFRS EWSDLLSDLQ KAGKITDEEA TKSSYLNIVG LVGSIDNDFC GTDMTIGTDS ALHRIMEIVD AITTTAQSHQ RTFVLEVMGR HCGYLALVTS LSCGADWVFI PECPPDDDWE EHLCRRLSET RTRGSRLNII IVAEGAIDKN GKPITSEDIK NGSRMGVEAV MALLEGTPDT PACVVSLSGN QAVRLPLMEC VQVTKDVTKA MDEKKFDEAL KLRGRSFMNN WEVYKLLAHV RPPVSKSGSH TVAVMNVGAP AAGMNAAVRS TVRIGLIQGN RVLVVHDGFE GLAKGQIEEA GWSYVGGWTG QGGSKLGTKR TLPKKSFEQI SANITKFNIQ GLVIIGGFEA YTGGLELMEG RKQFDELCIP FVVIPATVSN NVPGSDFSVG ADTALNTICT TCDRIKQSAA GTKRRVFIIE TMGGYCGYLA TMAGLAAGAD AAYIFEEPFT IRDLQANVEH LVQKMKTTVK RGLVLRNEKC NENYTTDFIF NLYSEEGKGI FDSRKNVLGH MQQGGSPTPF DRNFATKMGA KAMNWMSGKI KESYRNGRIF ANTPDSGCVL GMRKRALVFQ PVAELKDQTD FEHRIPKEQW WLKLRPILKI LAKYEIDLDT SDHAHLEHIT RKRSGEAAV), shown in SEQ ID NO.3 (MHKDEFHLKF FMCVIQSRQL VRTPQRTAGE ASTSSMLIPK PPPKTDILKS LDTMDDPDTV GSIPVFKTEW IMTHEEHHAA KTLGIGKAIA VLTSGGDAQG MNAAVRAVVR VGIFTGARVF FVHEGYQGLV DGGDHIKEAT WESVSMMLQL GGTVIGSARC KDFREREGRL RAAYNLVKRG ITNLCVIGGD GSLTGADTFR SEWSDLLSDL QKAGKITDEE ATKSSYLNIV GLVGSIDNDF CGTDMTIGTD SALHRIMEIV DAITTTAQSH QRTFVLEVMG RHCGYLALVT SLSCGADWVF IPECPPDDDW EEHLCRRLSE TRTRGSRLNI IIVAEGAIDK NGKPITSEDI KNLVVKRLGY DTRVTVLGHV QRGGTPSAFD RILGSRMGVE AVMALLEGTP DTPACVVSLS GNQAVRLPLM ECVQVTKDVT KAMDEKKFDE ALKLRGRSFM NNWEVYKLLA HVRPPVSKSG SHTVAVMNVG APAAGMNAAV RSTVRIGLIQ GNRVLVVHDG FEGLAKGQIE EAGWSYVGGW TGQGGSKLGT KRTLPKKSFE QISANITKFN IQGLVIIGGF EAYTGGLELM EGRKQFDELC IPFVVIPATV SNNVPGSDFS VGADTALNTI CTTCDRIKQS AAGTKRRVFI IETMGGYCGY LATMAGLAAG ADAAYIFEEP FTIRDLQANV EHLVQKMKTT VKRGLVLRNE KCNENYTTDF IFNLYSEEGK GIFDSRKNVL GHMQQGGSPT PFDRNFATKM GAKAMNWMSG KIKESYRNGR IFANTPDSGC VLGMRKRALV FQPVAELKDQ TDFEHRIPKE QWWLKLRPIL KILAKYEIDL DTSDHAHLEH ITRKRSGEAA); or an amino acid sequence which is derived from the amino acid sequence shown in SEQ ID NO 1-3 and has the same function as the amino acid sequence shown in SEQ ID NO 1-3.
Preferably, the preparation is a kit for detecting muscle-derived phosphofructokinase 6 and polypeptide fragments thereof in urine or urine exosomes of diabetics.
Preferably, the kit comprises one or more of an immunization method of antigen-antibody reaction and a kit thereof, such as an aptamer antibody or an antibody fragment capable of specifically binding to muscle-derived phosphofructokinase 6 and polypeptide fragments thereof.
Preferably, the detection method comprises a method for directly detecting mass spectrum of the muscle-source 6 phosphofructokinase and polypeptide fragments thereof and the like and a related kit thereof.
Preferably, the detection method comprises methods such as related nucleic acid detection for directly detecting muscle-derived phosphofructokinase 6 and polypeptide fragments thereof and related kits thereof.
Preferably, the kit further comprises a component selected from the group consisting of: solid phase carrier, diluent, reference substance, standard substance, quality control substance, detection antibody, secondary antibody diluent, luminous reagent, washing liquid, color developing liquid and stop solution.
Preferably, the standard comprises a muscle-derived phosphofructokinase 6 standard, a humanized tag antibody standard; preferably, the quality control product comprises: muscle source 6 phosphofructokinase quality control and humanized tag antibody quality control; preferably, the solid support comprises: microparticles, microspheres, slides, test strips, plastic beads, liquid phase chips, microwell plates or affinity membranes, and other carriers of equivalent function.
Preferably, the solid phase carrier is made of any one of polyvinyl chloride, polystyrene, polyacrylamide and cellulose and has similar functions.
The inventors first collected urine samples from healthy and diabetic patients, extracted urine exosomes by reference to human urine proteome program (Human Urine Proteome Project, HKUPP) and european kidney and urine proteome (European Kidney and Urine Proteomics, euroKUP) standard procedures, identified urine exosomes by transmission electron microscopy, exosome nanoparticle tracking detection and western immunoblotting, processed by liquid phase secondary mass spectrometry (liquid chromatography tandem mass spectrometry, LC-MS/MS) using Data independent scanning mode (Data-independent acquisition, DIA), and processed by quantitative value correction/calculation of absolute quantitative values, differential analysis and global cluster analysis of target proteins. And (3) quantitatively calculating data obtained in mass spectra of the diabetes group and the normal control group, and finally obtaining the phosphofructokinase of the differential protein muscle source 6.
Compared with healthy people, the invention proves that the muscle-source 6 phosphofructokinase and the polypeptide fragment thereof in urine or urine exosomes are in low expression in diabetics, and have better consistency with clinical diagnosis. Therefore, the application of the muscle-source 6 phosphofructokinase and the polypeptide fragment thereof in the diagnosis, differential diagnosis, disease degree judgment, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like of diabetes is proposed.
The invention plays the advantages of noninvasive acquisition of urine specimen, large-scale repeated sampling and convenient preservation, and utilizes the urine specimen to detect the muscle-source 6 phosphofructokinase and the polypeptide fragments thereof in urine or urine exosomes.
The foregoing and other objects, features and advantages of the invention will be apparent from the following more particular description of preferred embodiments, as illustrated in the accompanying drawings.
Drawings
FIG. 1 is a graph showing the content of muscle-derived phosphofructokinase 6 and polypeptide fragments thereof in a diabetic group and a healthy control group in urine or urine exosomes.
FIG. 2 is a graph showing the results of ELISA assays of muscle-derived phosphofructokinase 6 and polypeptide fragments thereof in urine or urine exosomes in diabetic groups and healthy control groups.
Detailed Description
Example 1Urine specimen collection and processing
Diabetes patients are selected as diabetes groups, and synchronous healthy physical examination persons are selected as normal control groups. Collecting fresh morning urine samples of each group of study subjects after admission, and collecting urine in the morning catheter of the study subjects which cannot normally urinate, and placing the study subjects in a dry and clean container. Exosomes in concentrated urine samples were extracted, resuspended in pre-chilled PBS and stored in a-80℃refrigerator, with reference to standard procedures for HKUPP and EuroKUP (http:// www.eurokup.org). The urine exosomes are identified by transmission electron microscopy, exosome nanoparticle tracking detection and western blotting.
Example 2Urine exosome polypeptide mass spectrometry
And (3) carrying out protein extraction and enzymolysis desalination on the urine sample, and measuring the concentration of the extracted protein. By advanced liquid-phase secondary mass spectrometry (LC-MS/MS), a data independent scanning mode (DIA) is adopted to construct urine exosome protein maps of normal people with different ages and different sexes.
Example 3Differential polypeptide data analysis
All experiments were performed in triplicate and all values are expressed as mean ± standard deviation. Fold change is 1.5 times to obtain the analysis result of the differential protein,P<0.05 was considered statistically significant, and differential proteins were screened for global cluster analysis of target proteins.
Muscle-derived phosphofructokinase 6 in urine or urine exosomes was expressed in low levels in diabetic patients compared to healthy persons, and the levels in normal control and diabetic groups were shown in fig. 1, with significant differences in expression.
Example 4Verification of muscle-derived phosphofructokinase 6 in urine or urine exosomes
The results of the enzyme-linked immunosorbent assay (ELISA) for muscle-derived phosphofructokinase 6 in urine or urine exosomes of healthy people and diabetics are shown in FIG. 2. The content of the muscle-source 6 phosphofructokinase in urine or urine exosome is reduced in diabetics, and the difference has statistical significancep<0.01）。
Although the invention has been described with respect to the preferred embodiments, it will be understood by those skilled in the art that various changes and modifications may be made without departing from the spirit and scope of the invention, and the invention is defined in the following claims.
Sequence listing
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<120> urine muscle-derived 6-phosphofructokinase and application of polypeptide fragment thereof in diabetes
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Tyr Cys Gly Tyr Leu Ala Thr Met Ala Gly Leu Ala Ala Gly Ala Asp
545 550 555 560
Ala Ala Tyr Ile Phe Glu Glu Pro Phe Thr Ile Arg Asp Leu Gln Ala
565 570 575
Asn Val Glu His Leu Val Gln Lys Met Lys Thr Thr Val Lys Arg Gly
580 585 590
Leu Val Leu Arg Asn Glu Lys Cys Asn Glu Asn Tyr Thr Thr Asp Phe
595 600 605
Ile Phe Asn Leu Tyr Ser Glu Glu Gly Lys Gly Ile Phe Asp Ser Arg
610 615 620
Lys Asn Val Leu Gly His Met Gln Gln Gly Gly Ser Pro Thr Pro Phe
625 630 635 640
Asp Arg Asn Phe Ala Thr Lys Met Gly Ala Lys Ala Met Asn Trp Met
645 650 655
Ser Gly Lys Ile Lys Glu Ser Tyr Arg Asn Gly Arg Ile Phe Ala Asn
660 665 670
Thr Pro Asp Ser Gly Cys Val Leu Gly Met Arg Lys Arg Ala Leu Val
675 680 685
Phe Gln Pro Val Ala Glu Leu Lys Asp Gln Thr Asp Phe Glu His Arg
690 695 700
Ile Pro Lys Glu Gln Trp Trp Leu Lys Leu Arg Pro Ile Leu Lys Ile
705 710 715 720
Leu Ala Lys Tyr Glu Ile Asp Leu Asp Thr Ser Asp His Ala His Leu
725 730 735
Glu His Ile Thr Arg Lys Arg Ser Gly Glu Ala Ala Val
740 745
<210> 3
<211> 850
<212> PRT
<213> Human Urine
<400> 3
Met His Lys Asp Glu Phe His Leu Lys Phe Phe Met Cys Val Ile Gln
1 5 10 15
Ser Arg Gln Leu Val Arg Thr Pro Gln Arg Thr Ala Gly Glu Ala Ser
20 25 30
Thr Ser Ser Met Leu Ile Pro Lys Pro Pro Pro Lys Thr Asp Ile Leu
35 40 45
Lys Ser Leu Asp Thr Met Asp Asp Pro Asp Thr Val Gly Ser Ile Pro
50 55 60
Val Phe Lys Thr Glu Trp Ile Met Thr His Glu Glu His His Ala Ala
65 70 75 80
Lys Thr Leu Gly Ile Gly Lys Ala Ile Ala Val Leu Thr Ser Gly Gly
85 90 95
Asp Ala Gln Gly Met Asn Ala Ala Val Arg Ala Val Val Arg Val Gly
100 105 110
Ile Phe Thr Gly Ala Arg Val Phe Phe Val His Glu Gly Tyr Gln Gly
115 120 125
Leu Val Asp Gly Gly Asp His Ile Lys Glu Ala Thr Trp Glu Ser Val
130 135 140
Ser Met Met Leu Gln Leu Gly Gly Thr Val Ile Gly Ser Ala Arg Cys
145 150 155 160
Lys Asp Phe Arg Glu Arg Glu Gly Arg Leu Arg Ala Ala Tyr Asn Leu
165 170 175
Val Lys Arg Gly Ile Thr Asn Leu Cys Val Ile Gly Gly Asp Gly Ser
180 185 190
Leu Thr Gly Ala Asp Thr Phe Arg Ser Glu Trp Ser Asp Leu Leu Ser
195 200 205
Asp Leu Gln Lys Ala Gly Lys Ile Thr Asp Glu Glu Ala Thr Lys Ser
210 215 220
Ser Tyr Leu Asn Ile Val Gly Leu Val Gly Ser Ile Asp Asn Asp Phe
225 230 235 240
Cys Gly Thr Asp Met Thr Ile Gly Thr Asp Ser Ala Leu His Arg Ile
245 250 255
Met Glu Ile Val Asp Ala Ile Thr Thr Thr Ala Gln Ser His Gln Arg
260 265 270
Thr Phe Val Leu Glu Val Met Gly Arg His Cys Gly Tyr Leu Ala Leu
275 280 285
Val Thr Ser Leu Ser Cys Gly Ala Asp Trp Val Phe Ile Pro Glu Cys
290 295 300
Pro Pro Asp Asp Asp Trp Glu Glu His Leu Cys Arg Arg Leu Ser Glu
305 310 315 320
Thr Arg Thr Arg Gly Ser Arg Leu Asn Ile Ile Ile Val Ala Glu Gly
325 330 335
Ala Ile Asp Lys Asn Gly Lys Pro Ile Thr Ser Glu Asp Ile Lys Asn
340 345 350
Leu Val Val Lys Arg Leu Gly Tyr Asp Thr Arg Val Thr Val Leu Gly
355 360 365
His Val Gln Arg Gly Gly Thr Pro Ser Ala Phe Asp Arg Ile Leu Gly
370 375 380
Ser Arg Met Gly Val Glu Ala Val Met Ala Leu Leu Glu Gly Thr Pro
385 390 395 400
Asp Thr Pro Ala Cys Val Val Ser Leu Ser Gly Asn Gln Ala Val Arg
405 410 415
Leu Pro Leu Met Glu Cys Val Gln Val Thr Lys Asp Val Thr Lys Ala
420 425 430
Met Asp Glu Lys Lys Phe Asp Glu Ala Leu Lys Leu Arg Gly Arg Ser
435 440 445
Phe Met Asn Asn Trp Glu Val Tyr Lys Leu Leu Ala His Val Arg Pro
450 455 460
Pro Val Ser Lys Ser Gly Ser His Thr Val Ala Val Met Asn Val Gly
465 470 475 480
Ala Pro Ala Ala Gly Met Asn Ala Ala Val Arg Ser Thr Val Arg Ile
485 490 495
Gly Leu Ile Gln Gly Asn Arg Val Leu Val Val His Asp Gly Phe Glu
500 505 510
Gly Leu Ala Lys Gly Gln Ile Glu Glu Ala Gly Trp Ser Tyr Val Gly
515 520 525
Gly Trp Thr Gly Gln Gly Gly Ser Lys Leu Gly Thr Lys Arg Thr Leu
530 535 540
Pro Lys Lys Ser Phe Glu Gln Ile Ser Ala Asn Ile Thr Lys Phe Asn
545 550 555 560
Ile Gln Gly Leu Val Ile Ile Gly Gly Phe Glu Ala Tyr Thr Gly Gly
565 570 575
Leu Glu Leu Met Glu Gly Arg Lys Gln Phe Asp Glu Leu Cys Ile Pro
580 585 590
Phe Val Val Ile Pro Ala Thr Val Ser Asn Asn Val Pro Gly Ser Asp
595 600 605
Phe Ser Val Gly Ala Asp Thr Ala Leu Asn Thr Ile Cys Thr Thr Cys
610 615 620
Asp Arg Ile Lys Gln Ser Ala Ala Gly Thr Lys Arg Arg Val Phe Ile
625 630 635 640
Ile Glu Thr Met Gly Gly Tyr Cys Gly Tyr Leu Ala Thr Met Ala Gly
645 650 655
Leu Ala Ala Gly Ala Asp Ala Ala Tyr Ile Phe Glu Glu Pro Phe Thr
660 665 670
Ile Arg Asp Leu Gln Ala Asn Val Glu His Leu Val Gln Lys Met Lys
675 680 685
Thr Thr Val Lys Arg Gly Leu Val Leu Arg Asn Glu Lys Cys Asn Glu
690 695 700
Asn Tyr Thr Thr Asp Phe Ile Phe Asn Leu Tyr Ser Glu Glu Gly Lys
705 710 715 720
Gly Ile Phe Asp Ser Arg Lys Asn Val Leu Gly His Met Gln Gln Gly
725 730 735
Gly Ser Pro Thr Pro Phe Asp Arg Asn Phe Ala Thr Lys Met Gly Ala
740 745 750
Lys Ala Met Asn Trp Met Ser Gly Lys Ile Lys Glu Ser Tyr Arg Asn
755 760 765
Gly Arg Ile Phe Ala Asn Thr Pro Asp Ser Gly Cys Val Leu Gly Met
770 775 780
Arg Lys Arg Ala Leu Val Phe Gln Pro Val Ala Glu Leu Lys Asp Gln
785 790 795 800
Thr Asp Phe Glu His Arg Ile Pro Lys Glu Gln Trp Trp Leu Lys Leu
805 810 815
Arg Pro Ile Leu Lys Ile Leu Ala Lys Tyr Glu Ile Asp Leu Asp Thr
820 825 830
Ser Asp His Ala His Leu Glu His Ile Thr Arg Lys Arg Ser Gly Glu
835 840 845
Ala Ala
850

Claims (9)

1. The application of muscle-source 6 phosphofructokinase and polypeptide fragments thereof in urine or urine exosomes in preparing preparations for diagnosis, differential diagnosis, judgment of disease degree, treatment effect evaluation, monitoring, prognosis evaluation, mechanism research and the like of diabetes.
2. The use according to claim 1, wherein the amino acid sequence of the muscle-derived phosphofructokinase 6 in urine or urine exosomes is shown in SEQ ID No. 1-3; or an amino acid sequence which is derived from the amino acid sequence shown in SEQ ID NO 1-3 and has the same function as the amino acid sequence shown in SEQ ID NO 1-3.
3. The use according to claim 1, wherein the preparation is a kit for detecting muscle-derived phosphofructokinase 6 and polypeptide fragments thereof in urine or urine exosomes of diabetics.
4. The use according to claim 3, wherein the kit comprises one or more of an immunization method of antigen-antibody reaction and a kit thereof such as an aptamer antibody or antibody fragment capable of specifically binding to muscle-derived phosphofructokinase 6 and polypeptide fragments thereof.
5. The use according to claim 3, wherein the detection method comprises a method for directly detecting mass spectrum of muscle-derived phosphofructokinase 6 and polypeptide fragments thereof and the like and a related kit thereof.
6. The use according to claim 3, wherein the detection method comprises a method for directly detecting muscle-derived phosphofructokinase 6 and its polypeptide fragment or its related nucleic acid detection, and its related kit.
7. The use according to claim 3, wherein the kit further comprises a component selected from the group consisting of: solid phase carrier, diluent, reference substance, standard substance, quality control substance, detection antibody, secondary antibody diluent, luminous reagent, washing liquid, color developing liquid and stop solution.
8. The use according to claim 7, wherein the standard comprises a muscle-derived phosphofructokinase 6 standard, a humanized tag antibody standard; preferably, the quality control product comprises: muscle source 6 phosphofructokinase control and humanized tag antibody quality control; preferably, the solid support comprises: microparticles, microspheres, slides, test strips, plastic beads, liquid phase chips, microwell plates or affinity membranes, and other carriers of equivalent function.
9. The use according to claim 8, wherein the solid support is made of any one of polyvinyl chloride, polystyrene, polyacrylamide, cellulose and a support with similar function.
CN202210429495.4A 2022-04-22 2022-04-22 Application of muscle-source 6 phosphofructokinase and polypeptide fragment thereof in urine or urine exosomes in diabetes Pending CN117147866A (en)

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