CN112111003A - 含有me序列的一类新型抑制血小板聚集和抗血栓形成寡肽 - Google Patents
含有me序列的一类新型抑制血小板聚集和抗血栓形成寡肽 Download PDFInfo
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- CN112111003A CN112111003A CN201910538828.5A CN201910538828A CN112111003A CN 112111003 A CN112111003 A CN 112111003A CN 201910538828 A CN201910538828 A CN 201910538828A CN 112111003 A CN112111003 A CN 112111003A
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Abstract
本发明提出了一种分离的寡肽。该分离的寡肽包括ME,所述ME为与Gαq蛋白相互作用位点。发明人首次从鱼皮胶原蛋白中分离鉴定出,其ME位点为与Gαq蛋白相互作用的活性位点,进而发明人通过实验,筛选出了包括ME的系列寡肽,ME为与Gαq蛋白相互作用位点。根据本发明实施例的系列寡肽能够与Gαq蛋白相互作用,抑制凝血酶的活性和血小板聚集,并且该系列寡肽具有耐胃肠道消化酶酶解和易吸收的特点,其在体内的稳定性增加。
Description
技术领域
本发明涉及生物医药领域,具体地,本发明涉及含有ME序列的一类新型抑制血小板聚集和抗血栓形成寡肽,更具体地,本发明涉及分离的寡肽,分离的核酸、构建体、重组细胞、药物组合物或保健品、寡肽在制备药物、保健品或试剂盒中的用途。
背景技术
血小板是哺乳动物血液中的有形成分之一,一般呈圆盘状、无细胞核,却具有丰富的胞内颗粒。在正常的生理条件下,血小板的主要功能是促进止血和加速凝血,同时还有维护毛细血管壁完整性的功能。但是血小板的过度活化参与许多病理性过程,包括血栓形成、动脉粥样硬化和癌症等(Blood Reviews,2005,19(2),111-123)。当受到激动剂刺激而活化时,血小板一方面被激活发生聚集,参与血栓形成;另一方面,活化的血小板发生释放反应,将贮存在致密颗粒、α-颗粒或溶酶体内的物质排出,释放的多种生物活性物质对血栓形成、动脉粥样硬化和癌症等的发生和发展具有非常重要的作用。从致密颗粒中释放的活性成分的功能是募集其他血小板发生聚集。从致密体释放的物质主要有腺苷二磷酸(ADP)、三磷酸腺苷(ATP)、5-羟色胺(5-HT)和Ca2+。其中ADP、5-HT和Ca2+均可促进血小板聚集,从而放大血小板的聚集作用。而α-颗粒含有大量的蛋白,这些蛋白包括粘着蛋白、趋化因子、细胞有丝分裂原和蛋白酶抑制剂。从血小板α-颗粒释放的细胞有丝分裂原含有多种生长因子,如TGF-β,ECGF,EGF,VEGF/VPF、IGF、bFGF和HGF等,这些生长因子也是重要的血管生成因子,促进肿瘤的发生和转移。而从α-颗粒中释放的趋化因和炎症因子,如IL-1β,CD40配体,PF4,MIP-1α和PDGF等可促进动脉粥样硬化的发生和发展(Trends in CardiovascularMedicine,2004,14(1),18-22.)。因此,抑制血小板的聚集和释放功能对预防和治疗血栓形成、动脉粥样硬化等心脑血管疾病和肿瘤的发生发展都具有重要意义。
抗血小板药物阿司匹林、氯吡格雷、噻氯匹定、阿昔单抗等在抑制血小板聚集以及抗血栓形成方面发挥重要作用,这些药物对于防治动脉粥样硬化和降低肿瘤发生发展的研究也广泛报道(中国循环杂志,2005,20(6),463-467;Blood 2018,131(16):1777–1789)。但是,这些药物的安全性、有效性使其临床应用受到一定的限制。因此,寻找和研究开发新的高效低毒的抗血小板活性成分具有十分重要的意义。
越来越多的研究表明来自天然食物或动植物的生物活性肽对人体健康有着积极的作用。这些生物活性肽能够通过调节人体内各种生理过程从而对人体产生有益影响,主要包括抗氧化肽、降血压肽、免疫调剂肽、抗菌肽等。近年来,抗血小板肽的研究得到越来越多的关注,相比传统的抗血小板药物,这些抗血小板活性肽具有毒副作用小、易吸收、药效持续时间长等优点。
发明内容
本申请是基于发明人对以下事实和问题的发现和认识作出的:
发明人通过大量的活性筛选,首次从鱼皮胶原蛋白中分离鉴定了二肽ME对凝血酶诱导的血小板聚集有特异性抑制作用,并进一步明确了其作用靶点是Gαq蛋白。发明人将筛选得到的ME二肽,并且通过氨基酸修饰得到了系列衍生物三肽,将含有ME序列的三肽与靶蛋白进行分子对接,虚拟筛选出了活性更高的Gαq蛋白小分子配体。根据细胞水平的高通量筛选和功能验证,开发出了具有抗血小板治疗前景的含ME序列的三肽。本发明所涉及的ME二肽和新型三肽序列结构,迄今尚未见相关报道。
为此,在本发明的第一方面,本发明提出了一种分离的寡肽。根据本发明的实施例,所述分离的寡肽包括ME,所述ME为与Gαq蛋白相互作用位点。发明人首次从鱼皮胶原蛋白中分离鉴定出,其ME位点为与Gαq蛋白相互作用的活性位点,进而发明人通过实验,筛选出了包括ME的系列寡肽,ME为与Gαq蛋白相互作用位点。根据本发明实施例的系列寡肽能够与Gαq蛋白相互作用,抑制凝血酶的活性和血小板聚集,并且发明人发现,该系列寡肽具有耐胃肠道消化酶酶解和易吸收的特点,其在体内的稳定性增加。
根据本发明的实施例,所述寡肽还可以进一步具有如下附加技术特征至少之一:
根据本发明的实施例,所述寡肽的长度为2~10个氨基酸。
根据本发明的实施例,所述寡肽的氨基酸序列如ME、HME、OME、WME或MET所示。发明人发现,具有上述序列的寡肽,其对血小板聚集的抑制作用更强。
在本发明的第二方面,本发明提出了一种分离的核酸。根据本发明的实施例,所述核酸编码前面所述的分离的寡肽。根据本发明实施例的核酸在导入受体细胞后,在适于蛋白表达的环境下,表达前面所述的分离的寡肽。所述寡肽具有Gαq蛋白相互作用位点,能够抑制Gαq蛋白,进而有效抑制凝血酶的活性和血小板聚集。
根据本发明的实施例,上述分离的核酸还可以进一步包括如下附加技术特征至少之一:
根据本发明的实施例,所述核酸具有SEQ ID NO:1~5任一项所示的核苷酸序列。
ATGGAG(SEQ ID NO:1),
CACATGGAG(SEQ ID NO:2),
CCCATGGAG(SEQ ID NO:3),
TGGATGGAG(SEQ ID NO:4),
ATGGAGACC(SEQ ID NO:5)。
根据本发明的实施例,SEQ ID NO:1所示的核苷酸序列编码ME,SEQ ID NO:2所示的核苷酸序列编码HME,SEQ ID NO:3所示的核苷酸序列编码OME,SEQ ID NO:4所示的核苷酸序列编码WME,SEQ ID NO:5所示的核苷酸序列编码MET。
在本发明的第三方面,本发明提出了一种构建体。根据本发明的实施例,所述构建体携带前面所述的核酸。根据本发明实施例的构建体导入细胞后,在适于蛋白表达的环境下,表达前面所述的分离的寡肽。所述寡肽具有Gαq蛋白相互作用位点,能够抑制Gαq蛋白,进而有效抑制凝血酶的活性和血小板聚集。
在本发明的第四方面,本发明提出了一种重组细胞。根据本发明的实施例,所述重组细胞携带前面所述的核酸或前面所述的构建体,进而在适于蛋白表达的环境下,表达前面所述的寡肽。
在本发明的第五方面,本发明提出了一种药物组合物或保健品。根据本发明的实施例,所述药物组合物包括前面所述的分离的寡肽、前面所述的分离的核酸或前面所述的构建体或前面所述的重组细胞。根据本发明实施例的药物组合或保健品,可有效抑制血小板聚集,对血栓、动脉粥样硬化或癌症疾病具有预防或治疗作用。
根据本发明的实施例,所述药物组合物或保健品还可以进一步包括如下附加技术特征至少之一:
根据本发明的实施例,所述药物组合物或保健品进一步包括可接受的辅料。根据本发明的具体实施例,所述物组合物或保健品可呈水性悬浮、溶液或固体状态。
在本发明的第六方面,本发明提出了前面所述的分离的寡肽、前面所述的分离的核酸或前面所述的构建体或前面所述的重组细胞在制备药物、保健品中的用途,所述药物或保健品用于抑制血小板聚集、治疗或预防Gαq蛋白相关性疾病。发明人发现,本申请的寡肽具有与Gαq蛋白相互作用的活性位点ME,可有效结合Gαq蛋白并抑制Gαq蛋白的活性。根据本发明实施例的药物或保健品以前面所述的多肽或表达前面所述多肽的核酸、构建体或重组细胞为活性成分,对Gαq蛋白相关性疾病具有有效的预防或治疗作用。
根据本发明的实施例,上述用途还可以进一步包括如下附加技术特征至少之一:
根据本发明的实施例,所述Gαq蛋白相关性疾病为血栓、动脉粥样硬化或癌症。发明人发现,根据本申请实施例的药物或保健品对血栓、动脉粥样硬化或癌症的治疗效果显著。
在本发明的第七方面,本发明提出了前面所述的分离的寡肽、前面所述的分离的核酸或前面所述的构建体或前面所述的重组细胞在制备试剂盒中的用途,所述试剂盒用于抑制Gαq蛋白或Gαq蛋白下游信号通路。如前所述,本申请的ME系列寡肽具有与Gαq蛋白相互作用的活性位点,可有效结合Gαq蛋白并抑制Gαq蛋白的活性。包含本申请实施例的多肽或表达前面所述多肽的核酸、构建体或重组细胞的试剂盒,在科学研究中,可有效用于抑制Gαq蛋白或Gαq蛋白下游信号通路。
附图说明
图1是根据本发明实施例的二肽ME及ME系列三肽的质谱图;
图2是根据本发明实施例的二肽ME及ME系列三肽对凝血酶诱导的血小板聚集的抑制作用结果图;
图3是根据本发明实施例的不同浓度二肽ME对血小板颗粒释放的影响的结果图;
图4是根据本发明实施例的二肽ME对血小板胞内Ca2+释放的影响的结果图;
图5是根据本发明实施例的二肽ME的作用靶点及分子机制图;
图6是根据本发明实施例的ME在模拟胃消化和肠消化过程中的HPLC色谱图;
图7是根据本发明实施例的二肽ME在人源血浆中的稳定性的结果图;
图8是根据本发明实施例的二肽ME对小鼠急性肺血栓形成的影响的结果图;
图9是根据本发明实施例的二肽ME的对凝血时间影响的结果图;
图10是根据本发明实施例的二肽ME对SD大鼠颈动脉血栓形成影响的结果图;
图11是根据本发明实施例的三肽WME对小鼠急性肺血栓形成的影响的结果图;
图12是根据本发明实施例的三肽WME的对凝血时间影响的结果图;以及
图13是根据本发明实施例的三肽WME对SD大鼠颈动脉血栓形成的影响的结果图。
具体实施方式
下面详细描述本发明的实施例,所述实施例的示例在附图中示出。下面通过参考附图描述的实施例是示例性的,旨在用于解释本发明,而不能理解为对本发明的限制。
本发明的第一个目的是提供具有抑制血小板聚集和抗血栓形成的生物活性肽。
本发明是在前期对胶原生物活性肽的抗血小板聚集活性评价基础上,筛选的具有高血小板聚集抑制率的小肽。相较传统的抗血小板药物,生物活性肽具有毒副作用小、更加安全的优势。并且本发明涉及的ME或一类具有ME结构序列的三肽具有耐胃肠道消化酶酶解和易吸收的特点。因此,本发明将对抗血小板聚集和抗血栓形成新药物和保健食品的开发具有重要意义。
本发明提供一类合成寡肽,所述的氨基酸结构序列分别为Met-Glu(ME)、His-Met-Glu(HME)、Hyp-Met-Glu(OME)、Trp-Met-Glu(WME)和Met-Glu-Thr(MET)。
本发明的第二个目的是提供上述ME及含ME序列的三肽在制备抗血小板聚集和血栓形成药物或保健食品中的应用。一种抗血小板聚集的药物或保健食品,含有上述抗血小板聚集的寡肽作为活性成分。
本发明的第三个目的是提供上述二肽ME及含ME序列的三肽在制备、预防和治疗血栓形成和动脉粥样硬化等疾病的药物或保健食品中的应用。
一种预防和治疗血栓形成和动脉粥样硬化等疾病的药物或保健食品,其含有上述抗血小板聚集的二肽ME或ME系列三肽作为活性成分。
下面结合具体实施例对本发明做进一步的说明,但下述实施例绝非对本发明有任何限制。
实施例1
实施例1中的合成多肽序列为Met-Glu、His-Met-Glu、Hyp-Met-Glu、Trp-Met-Glu和Met-Glu-Thr,由“浙江鸿拓科技有限公司”合成,通过质谱分析验证了合成物的纯度大于98%。结果由图1所示。
实施例2二肽ME及含ME序列的三肽对凝血酶诱导的血小板聚集的影响
(1)血小板的制备
Sprague Dawley(SD)大鼠(280-310g)腹腔注射戊巴比妥钠进行麻醉,戊巴比妥钠浓度2%(w/v),注射剂量50mg/Kg体重。待大鼠完全麻醉后进行腹主动脉取血。将1份3.8%的柠檬酸钠与9份全血混合抗凝,加入等体积的PBS混合。50g离心10min,取上层富血小板血浆(PRP)。PRP 750g离心10min,上层为贫血小板血浆(PPP),下层为血小板沉淀。血小板沉淀用HEPES/Tyrode’s缓冲液(10mM HEPES/NaOH,pH 7.4,5.56mMglucose,137mM NaCl,12mMNaHCO3,2.7mM KCl,0.36mM NaH2PO4,1mM MgCl2)清洗一次,再次750g离心10min,取下层血小板沉淀,悬浮于HEPES/Tyrode’s缓冲液,调整血小板浓度为2-3×108个/mL,得到洗涤血小板,备用。
(2)ME及ME系列三肽的血小板聚集抑制率
血小板聚集率的测定采用普利生LBY-NJ4型血小板聚集仪。将血小板聚集仪预热30min,将330μL HEPES/Tyrode’s缓冲液加入比色杯中,并用此调零。对于模型组和样品组,将270μL血小板预先在37℃孵育5min,然后加入30μL HEPES/Tyrode’s缓冲液(模型组)或样品(终浓度为2mM),37℃孵育5min。最后加入凝血酶(0.5U/mL)诱导血小板聚集,测定血小板在5min时的聚集率。5条活性肽对凝血酶诱导的血小板聚集率的影响如图2所示,ME系列三肽可以显著抑制凝血酶诱导的血小板聚集,其半数抑制率(IC50)由表1所示。由结果可以看到,ME系列三肽对凝血酶诱导的血小板聚集均具有较好的抑制作用。其中ME和WME的IC50分别为2.396mM和2.013mM。
表1:
实施例3以ME为例,说明含ME序列的系列寡肽抗血小板的药理作用及药代动力学实验
(1)二肽ME对血小板颗粒物释放的影响
参照实施例2中的方法制备洗涤血小板,取180μL血小板预先在37℃孵育5min,然后加入20μL HEPES/Tyrode’s缓冲液(模型组)或样品(终浓度为2mM),37℃孵育5min。最后加入凝血酶(0.5U/mL)诱导血小板激活,反应5min。反应结束后置于冰上1min终止反应,然后10000g离心2min,取上清液备用。对上清液中的β-血小板球蛋白(β-TG,血小板α颗粒释放标记物)和血清素(5-HT,血小板致密颗粒释放标记物)进行ELISA分析。二肽ME对血小板颗粒物释放的影响如图3所示,实验结果表明,二肽ME剂量依赖的抑制血小板α颗粒β-TG和血小板致密颗粒5-HT的释放,并且对致密颗粒释放的抑制效果强于对α颗粒释放的抑制效果。
(2)二肽ME对血小板胞内游离Ca2+浓度的影响
采用双波长荧光比色法测定Ca2+浓度。参照实施例2方法设置正常组、样品组和模型组,避光37℃孵育5min,加入凝血酶诱导5min,进行荧光检测。激发波长分别固定为340nm和380nm,波宽5nm,发射波长固定在510nm,波宽5nm,记录各组在激发波长340nm和380nm处测得的荧光强度比值R(F340/F380),然后加入Triton X-100(终浓度0.1%),测得Rmax,再加入EGTA(终浓度3mM)测定Rmin,根据如下公式计算血小板胞内Ca2+浓度。
[Ca2+]i=Kd×(R-Rmin)/(Rmax-R)×SFB
式中Kd为Fura-2/AM与Ca2+反应的解离常数,为224nmol/L,R为各测定点F340与F380荧光强度的比值。SFB为组成Rmin和Rmax的F380之间的比值。其测定结果如图4所示,二肽ME在一定浓度下可抑制血小板内钙离子动员。
(3)二肽ME的靶点验证及分子作用机制
参照实施例2方法制备洗涤血小板,利用磷酸化抗体芯片筛选潜在靶标,对表达差异蛋白进行Western Blot验证。初步确定ME作用的靶蛋白为Gαq。进一步的靶点验证实验如下。
取270μL的洗涤血小板于测试杯中,用不同浓度的A4、U73122或台式液分别置于血小板聚集仪37℃孵育4min,转入测试通道,加入凝血酶(0.5U/mL)诱导血小板聚集,测定3min内的血小板聚集率。同时,以YM254890为阳性对照,设置未经抑制剂和激动剂处理的血小板为正常组。将经抑制剂和A4处理的血小板750×g离心1min,收集血小板沉淀,加入RIPA裂解液(含蛋白酶抑制剂),于冰上裂解20min,裂解结束后4℃下13000×g离心10min,取上清液,立即使用BCA试剂盒进行蛋白浓度测定。将各样品调整浓度后加入6×上样缓冲液混合,沸水浴3min,分装,-20℃保存,用于免疫印迹分析。其结果如图5所示,二肽ME可通过抑制Gαq抑制凝血酶诱导的血小板激活,进而抑制下游信号的转导和血小板的活化。
(4)二肽ME的胃肠耐受性实验
将二肽ME溶解于去离子水中,用盐酸调节体系pH为2.0,加入胃蛋白酶在37℃下消化2h,胃消化结束后用0.9M的NaHCO3将胃消化物pH调节至5.3,再加入1M NaOH调节pH至7.5,然后加入胰酶在37℃进一步消化4h,消化结束后沸水浴10min灭活,待冷却至室温,调节pH至7.0,离心浓缩冷冻干燥,得到模拟胃肠消化物,用于HPLC分析。其结果如图6所示,ME在胃消化和肠消化结束后其色谱峰的保留时间和峰面积无显著变化,说明没有被降解,对胃蛋白酶和胰蛋白酶具有耐受性。
(5)二肽ME的血浆稳定性实验
利用真空采血管采集人体肘静脉血液10mL,10000g下4℃离心10min,取上清血浆备用;配制10mM的二肽ME,按照1:20(v/v)加入到血浆中,使肽的终浓度为500μM,37℃下水浴振荡,分别在0、0.5、1、2、4、8h取样100μL,经3kDa超滤离心管在14000g离心15min,收集滤出液进行HPLC检测多肽含量。血浆稳定性及半衰期如图7所示,随着孵育时间的延长,ME在血浆中的保留量降低,半衰期约为2h。
(6)二肽ME的凝血级联反应
SD大鼠腹腔注射戊巴比妥钠进行麻醉,然后腹主动脉取血,将1份3.8%的柠檬酸钠与9份全血混合,1500g离心15min,取上层血浆,备用。取180μL血浆与20μL胶原肽混合,使胶原肽终浓度为2mM,同时以生理盐水为阴性对照,以0.5mM的Agatroban(阿加曲班)为阳性对照,37℃孵育5min,结束后立即用全自动血凝仪测定PT、APTT和TT。其结果如表2所示,经阳性对照阿加曲班处理后,PT、APTT和TT显著增加,均超过120秒。ME处理后PT、APTT和TT没有显著变化,表明ME对内源和外源凝血途径没有显著影响,也不会抑制凝血酶活性。
表2:二肽ME对凝血反应的影响
注:PT,凝血酶原时间;APTT,活化部分凝血活酶时间;TT,凝血酶时间
实施例4二肽ME对KM小鼠急性肺血栓形成的影响
对雄性KM小鼠进行适应性饲养6天,随后将小鼠分为4组,每组10只,分别为生理盐水组、ME低剂量组(150μM/kg bw.)、ME高剂量组(300μM/kg bw.)、氯吡格雷组(45mg/kgbw.)。进行灌胃,2h后进行急性肺血栓诱导,所有组别尾静脉注射胶原、肾上腺素混合溶液造模(5%葡萄糖配制375μg/ml胶原和14.3μg/ml肾上腺素混合注射液,0.05mL/10g),观察小鼠生理反应、记录死亡时间和恢复时间,计算10min内小鼠存活率。结果如图8所示,经二肽ME灌胃2h后小鼠存活率接近80%,具有良好的抗血栓功效。
实施例5二肽ME对KM小鼠断尾凝血时间的影响
参照实施例4的方法对KM小鼠进行分组,每组9只进行灌胃,2h后剪断小鼠尾尖3mm,每15s用滤纸片检查小鼠尾部出血情况,直至没有出血,记录凝血时间。结果如图9所示,阳性对照氯吡格雷显著延长小鼠断尾后的凝血时间,凝血时间超过15min,与空白对照组相比,二肽ME对小鼠凝血功能没有影响,没有出血风险。
实施例6二肽ME对SD大鼠颈动脉血栓形成的影响
对雄性SD大鼠进行适应性饲养6天,随后将大鼠分为4组,每组5只,分别为生理盐水组、ME低剂量组(100μM/kg bw.)、ME高剂量组(200μM/kg bw.)、氯吡格雷组(30mg/kgbw.)。用2%戊巴比妥钠(0.25mL/100g)对SD大鼠麻醉后,沿颈正中线切开,钝性分离右侧1.5cm长颈动脉,放入1cm宽的封口胶条,浸有10%FeCl3溶液的滤纸条(1cm×0.5cm)环裹分离备用的颈动脉段,并用封口胶条封住15min;40min后,结扎滤纸条两段血管(或者拿止血钳夹住两段血管),精确剪下滤纸条包裹的血管段,用洁净滤纸吸干血管内余血,精确称量含血栓的血管湿重,取出血栓后的血管再称重,两者相减即为该1cm长血管段内血栓的质量。结果如图10所示,二肽ME可有效减少颈动脉血栓形成的重量,具有抗血栓形成作用。
实施例7三肽WME对KM小鼠急性肺血栓形成的影响
对雄性KM小鼠进行适应性饲养6天,随后将小鼠分为4组,每组10只,分别为生理盐水组、WME低剂量组(150μM/kg bw.)、WME高剂量组(300μM/kg bw.)、氯吡格雷组(45mg/kgbw.)。急性肺血栓诱导方法参照实施例4。结果如图11所示,经三肽WME灌胃2h后小鼠存活率相比空白对照组显著提高,具有良好的抗血栓功效。
实施例8三肽WME对KM小鼠断尾凝血时间的影响
参照实施例7的方法对KM小鼠进行分组,每组9只进行灌胃,2h后剪断小鼠尾尖3mm,每15s用滤纸片检查小鼠尾部出血情况,直至没有出血,记录出血时间。结果如图12所示,三肽WME对小鼠凝血功能没有影响,没有出血风险。
实施例9三肽WME对SD大鼠颈动脉血栓形成的影响
对雄性SD大鼠进行适应性饲养6天,随后将大鼠分为4组,每组5只,分别为生理盐水组、WME低剂量组(100μM/kg bw.)、WME高剂量组(200μM/kg bw.)、氯吡格雷组(30mg/kgbw.)。大鼠颈动脉血栓实验方法参照实施例6。计算血栓重量,结果如图13所示,三肽WME可有效减少颈动脉血栓形成的重量,具有抗血栓形成作用。
在本说明书的描述中,参考术语“一个实施例”、“一些实施例”、“示例”、“具体示例”、或“一些示例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或者特点包含于本发明的至少一个实施例或示例中。在本说明书中,对上述术语的示意性表述不必须针对的是相同的实施例或示例。而且,描述的具体特征、结构、材料或者特点可以在任一个或多个实施例或示例中以合适的方式结合。此外,在不相互矛盾的情况下,本领域的技术人员可以将本说明书中描述的不同实施例或示例以及不同实施例或示例的特征进行结合和组合。
尽管上面已经示出和描述了本发明的实施例,可以理解的是,上述实施例是示例性的,不能理解为对本发明的限制,本领域的普通技术人员在本发明的范围内可以对上述实施例进行变化、修改、替换和变型。
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Claims (12)
1.一种分离的寡肽,其特征在于,包括ME,所述ME为与Gαq蛋白相互作用位点。
2.根据权利要求1所述分离的寡肽,其特征在于,所述寡肽的长度为2~10个氨基酸。
3.根据权利要求1所述分离的寡肽,其特征在于,所述寡肽的氨基酸序列如ME、HME、OME、WME或MET所示。
4.一种分离的核酸,其特征在于,编码权利要求1~3任一项所述的分离的寡肽。
5.根据权利要求4所述的分离的核酸,其特征在于,所述核酸具有SEQ ID NO:1~5任一项所示的核苷酸序列。
6.一种构建体,其特征在于,携带权利要求4~5任一项所述的核酸。
7.一种重组细胞,其特征在于,携带权利要求4~5任一项所述的核酸或权利要求6所述的构建体。
8.一种药物组合物或保健品,其特征在于,包括权利要求1~3任一项所述的分离的寡肽、权利要求4~5任一项所述的分离的核酸或权利要求6所述的构建体或权利要求7所述的重组细胞。
9.根据权利要求8所述的药物组合物或保健品,其特征在于,进一步包括可接受的辅料。
10.权利要求1~3任一项所述的分离的寡肽、权利要求4~5任一项所述的分离的核酸或权利要求6所述的构建体或权利要求7所述的重组细胞在制备药物、保健品中的用途,所述药物或保健品用于抑制血小板聚集、治疗或预防Gαq蛋白相关性疾病。
11.根据权利要求10所述的用途,其特征在于,所述Gαq蛋白相关性疾病为血栓、动脉粥样硬化或癌症。
12.权利要求1~3任一项所述的分离的寡肽、权利要求4~5任一项所述的分离的核酸或权利要求6所述的构建体或权利要求7所述的重组细胞在制备试剂盒中的用途,所述试剂盒用于抑制Gαq蛋白或Gαq蛋白下游信号通路。
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