CN114163516B - 一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用 - Google Patents
一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用 Download PDFInfo
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Abstract
本发明公开了一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用，该制备方法包括以下步骤:(1)鱼皮胶原的提取；(2)限制性酶解；(3)固定化铜锌离子亲和层析；(4)液相色谱‑串联质谱联用鉴定；(5)分子对接筛选。筛选出4条与酪氨酸酶螯合最为紧密的肽，这4条肽的氨基酸序列分别为FSGLD、PFRMY、RGFTGM和APGAPGGGF。采用本发明方法制备的胶原蛋白源酪氨酸酶抑制肽具有良好的酪氨酸酶抑制活性，可作为美白功能成分用于化妆品中，具有良好的应用前景。
Description
技术领域
本发明属于动物源生物活性肽领域，具体涉及一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用。
背景技术
黑色素是决定人类肤色的关键因子，正常含量的黑色素可在皮肤表皮形成保护屏障抵抗紫外线的损伤，但过量沉积会导致皮肤问题如痤疮、黄褐斑、黑素病、脂溢性角化病等。酪氨酸酶(tyrosinase,TYR)是黑色素生物合成的关键酶，催化从酪氨酸生成多巴醌的限速反应。针对黑色素过量沉积问题，抑制酪氨酸酶的活性是一种有效方式。尽管关于新型酪氨酸酶抑制剂的报道较多，但抑制效果不甚理想，只有少数几种如曲酸、对苯二酚、熊果苷等具有较好的抑制效果，但由于细胞毒性和诱变性被限制使用。因此，需要开发没有毒性和长期稳定性的新型抑制剂。
食物蛋白源酪氨酸酶抑制肽具有分子量小、易吸收、生物相容性好、安全性高的特点，在保健品及化妆品领域具有广阔的应用前景。食物蛋白源酪氨酸酶抑制肽的来源广泛，近年来以水产动物蛋白源为研究热点，但以罗非鱼皮为原料制备酪氨酸酶抑制肽未见报道。罗非鱼(Oreochromismossambicus)是一种淡水经济鱼类，我国产量居世界第一。罗非鱼的加工以冷冻鱼片为主，剩余部位作为副产物被加工成饲料或直接丢弃。但罗非鱼的加工副产物中含有约15％的粗蛋白，且鱼皮中含有丰富的胶原蛋白，是一种优质蛋白资源。因此以罗非鱼皮为原料制备酪氨酸酶抑制肽一方面可提高罗非鱼产业的精深加工水平与高值化利用率，另一方面可降低环境对加工废弃物的承受压力。
酪氨酸酶的活性中心是以两个铜离子为催化位点的疏水空腔，肽可与活性中心的铜离子形成相互作用力，螯合越紧密，使得铜离子催化活性越低，肽的抑制活性就越高。另外，TRP-2是酪氨酸酶相关蛋白家族的主要蛋白之一，在黑色素合成路径中负责将多巴色素催化生成二羟基吲哚-羧酸，此过程需要锌离子的参与。具有锌离子螯合活性的肽可与TRP-2竞争锌离子，阻断黑色素的生成。本发明通过固定化铜锌离子亲和层析提取具有酪氨酸酶抑制活性的肽组分，简化了纯化过程，提取得到的肽其酪氨酸酶抑制活性与传统手段获得的肽活性一致。
大量实验研究表明，含有天冬氨酸、苯丙氨酸、缬氨酸、赖氨酸、精氨酸、谷氨酸、丝氨酸等的肽表现出较强的金属螯合能力，尤其是天冬氨酸、苯丙氨酸、精氨酸及丝氨酸对铜与锌均具有很好的螯合能力，大量研究表明这几种氨基酸对肽的酪氨酸酶抑制活性具有突出贡献，能帮助肽更易螯合酪氨酸酶的活性中心。将铜锌离子用于固定化亲和层析，可纯化得到富含天冬氨酸、苯丙氨酸、精氨酸及丝氨酸等特征性氨基酸的肽组分。相较于化学合成酪氨酸酶抑制剂，活性肽具备更安全、更易吸收、效果更佳的优势。
《具有酪氨酸酶抑制作用的活性肽》借助在线酶切工具对胶原蛋白肽序列进行虚拟酶切，通过Discovery Studio软件对所得肽序列进行分子对接筛选，得到三条最具潜在酪氨酸酶抑制活性的肽Ser-Asp-Trp、Gly-Ala-Arg、Asp-Gly-Leu，其IC50值分别为0.84mg/mL、0.34mg/mL及0.15mg/mL。该专利通过虚拟酶解筛选酪氨酸酶抑制肽，无法保证实际酶解操作可获得相同肽段，亦无法得知纯肽组分的抑制活性是否高于混合肽组分，仅适用于作为纯肽成分添加的产品，提高了生产成本。
发明内容
为了解决现有技术存在的不足，本发明提供了一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用。
本发明的目的通过以下技术方案来实现。
一种胶原蛋白源酪氨酸酶抑制肽，所述胶原蛋白源酪氨酸酶抑制肽的氨基酸序列为FSGLD、PFRMY、RGFTGM或APGAPGGGF。
所述胶原蛋白源酪氨酸酶抑制肽，其氨基酸序列分别为:FSGLD、PFRMY、RGFTGM和APGAPGGGF。五肽FSGLD、五肽PFRMY、六肽RGFTGM和九肽APGAPGGGF对酪氨酸酶活性的半抑制浓度(IC50)分别为0.279mg/mL、0.109mg/mL、0.377mg/mL和0.658mg/mL。
一种胶原蛋白源酪氨酸酶抑制肽的制备方法，包括以下步骤：
(1)鱼皮胶原的提取：取罗非鱼皮，加入NaOH溶液浸泡，浸泡后用流水洗至中性，再加入盐酸浸泡，用流水洗至中性后沥干打浆，将浆液置于恒温水浴，离心取上清液即得罗非鱼皮胶原提取液；
(2)限制性酶解：取罗非鱼皮胶原提取液，根据罗非鱼皮胶原提取液中的蛋白质质量加入蛋白酶，保温震荡反应，反应结束后沸水浴灭酶，抽滤后离心取上清液干燥，即得到罗非鱼皮酶解粉；
(3)固定化铜锌离子亲和层析：将罗非鱼皮酶解粉溶解在缓冲液中制成酶解液，通过固定化铜锌离子亲和层析柱，用洗脱液洗脱并收集纯化罗非鱼皮肽组分，冷冻干燥，得到纯化罗非鱼皮肽组分冻干样，即为具有铜锌离子螯合活性的罗非鱼皮肽；
(4)液相色谱-串联质谱联用鉴定：将纯化罗非鱼皮肽组分冻干样经由配备在线纳喷离子源的液相色谱-串联质谱联用(LC-MS/MS)分析，串联质谱图经过PEAKS Studioversion 10.6分析，所得罗非鱼皮肽序经De Novo检测及PEAKS搜库后选取Verifiable denovo sequences序列作为纯化罗非鱼皮肽最终组分；
(5)分子对接筛选：通过分子对接软件AutodockVina对所得罗非鱼皮肽序进行模拟对接，筛选出4条与酪氨酸酶螯合最为紧密的肽序列，并利用Pymol及Discovery Studio软件进行可视化处理。
进一步地，步骤(3)所述收集纯化罗非鱼皮肽组分的保留时间为38min。
进一步地，步骤(1)所述罗非鱼皮与NaOH溶液或盐酸的质量比为1:(5～10)；步骤(1)所述NaOH溶液的浓度为0.01～0.2mol/L，NaOH溶液浸泡的时间为20～30min；步骤(1)所述盐酸的浓度为0.01～0.2mol/L，盐酸浸泡的时间为10～30min；步骤(1)所述水浴的温度为50～65℃；步骤(1)所述水浴的时间为3～8h。
进一步地，步骤(2)所述蛋白酶为碱性蛋白酶、中性蛋白酶、胰蛋白酶、猪胰凝乳蛋白酶、木瓜蛋白酶中的任意一种；步骤(2)所述蛋白酶的添加量为蛋白质质量的0.5～2％。
进一步地，步骤(2)所述保温震荡反应的温度为25-55℃，所述保温震荡反应的时间为2～8h；步骤(2)所述灭酶的时间为15min；步骤(2)所述离心的转速为8000～10000r/min，离心的时间为15～30min。
进一步地，步骤(2)中蛋白酶的酶解时间优选为2～8h。
进一步地，步骤(3)所述固定化铜锌离子亲和层析柱用的硫酸铜溶液与硫酸锌溶液混合制备，其中硫酸铜溶液的浓度为0.2～0.6mol/L；步骤(3)所述硫酸锌与硫酸铜的物质的量之比为1:(1～3)。
进一步地，步骤(3)所述固定化铜锌离子亲和层析中，使用缓冲液的pH值为7.6，洗脱液是pH值为4.0且含NaCl浓度为0.5mol/L的溶液；步骤(3)所述固定化铜锌离子亲和层析中，样品上样量为5～10mg/次，螯合时间为40～90min，洗脱速度为1～5mL/min。
进一步地，步骤(4)所述液相色谱-串联质谱联用分析中，液相色谱仪上样体积为8～10μL，样品分离梯度为30～40min，柱流量为300～500nL/min，流动相的A相：体积百分比0.1％甲酸水溶液，流动相的B相：含体积百分比0.1％甲酸的乙腈(ACN)溶液，柱温为40℃，电喷雾电压2kV。
进一步地，步骤(4)所述液相色谱-串联质谱联用分析中，质谱仪(MS)参数设置如下：MS:质荷比(m/z)扫描范围＝100-1500；分辨率＝70000；离子捕获数量(AGC target)＝3e6；最大注入时间＝50ms；扫描电荷＝1-6；高能碰撞(HCD-MS/MS)(top 20)：分辨率＝17500；隔离窗口＝2质荷比；离子捕获数量＝1e5；最大注入时间＝45ms；碰撞能量＝28,动态排除时间＝30s。
本发明还提供了所述胶原蛋白源酪氨酸酶抑制肽在美白面霜中的应用。
本发明与现有技术相比，具有如下有益效果：
(1)以加工废料罗非鱼皮为原料，通过酶解与首次利用固定化铜锌离子亲和层析分离得到了能够有效抑制酪氨酸酶活性的五肽FSGLD、五肽PFRMY、六肽RGFTGM和九肽APGAPGGGF，在保证抑制效果同时减少纯化步骤，提高制备效率；使用两种金属离子筛选目标肽段，使获得的肽段具有更高的纯度。并明确了活性肽与酪氨酸酶螯合的作用机制，为酪氨酸酶抑制肽的制备提供了新途径，同时促进了低值水产品的高值化利用。
(2)本发明得到的4条活性肽通过螯合酪氨酸酶的活性中心抑制酶活性，从而减少黑色素的生成，因此可作为美白功能成分用于化妆品中，具有良好的应用前景。
(3)本发明使用两种金属离子筛选目标肽段，使获得的肽段具有更高的纯度。
附图说明
图1为五肽FSGLD与酪氨酸酶的分子对接图。
图2为五肽PFRMY与酪氨酸酶的分子对接图。
图3为六肽RGFTGM与酪氨酸酶的分子对接图。
图4为九肽APGAPGGGF与酪氨酸酶的分子对接图。
图5为固定化铜锌离子亲和层析色谱图。
具体实施方式
以下螯合具体实施例对本发明作进一步的说明。
实施例1
(1)取罗非鱼皮，加入0.01mol/L NaOH溶液，加入罗非鱼皮和NaOH溶液的质量比为1:5，浸泡20min后流水洗至中性；再加入0.01mol/L盐酸，加入罗非鱼皮和盐酸的质量比为1:5，浸泡10min后流水洗至中性，沥干打浆，将浆液置于50℃恒温水浴，3h后离心取上清液浓缩，得罗非鱼皮胶原提取液；
(2)取鱼皮胶原，加入碱性蛋白酶，碱性蛋白酶的添加量为罗非鱼皮胶原提取液中蛋白质质量的0.5％，在pH9.0，55℃条件下保温震荡反应2h，反应结束后沸水浴灭酶15min，抽滤，再以转速为8000r/min离心15min，取上清液干燥，得罗非鱼皮酶解粉；
(3)将罗非鱼皮酶解粉溶解在缓冲液中制成酶解液，以0.2mol/L硫酸铜溶液与0.2mol/L硫酸锌溶液等体积混合制备柱，将酶解液通过固定化铜锌离子亲和层析柱，上样量为5mg/次，螯合时间为40min，用pH值为7.6的缓冲液洗去未螯合组分，再用pH值为4.0且含NaCl浓度为0.5mol/L的溶液作为洗脱液洗脱螯合肽，洗脱速度为1mL/min，冷冻干燥，得到纯化罗非鱼皮肽组分冻干样，即为具有铜锌离子螯合活性的罗非鱼皮肽；
(4)将纯化罗非鱼皮肽组分冻干样经由配备在线纳喷离子源的液相色谱-串联质谱联用分析，液相色谱仪上样体积为8μL，样品分离梯度为30min，柱流量控制在300nL/min，流动相的A相：0.1％甲酸水溶液，B相：含0.1％甲酸的乙腈溶液，柱温为40℃，电喷雾电压2kV；质谱仪参数设置如下：MS:质荷比扫描范围＝100-1500；分辨率＝70000；离子捕获数量＝3e6；最大注入时间＝50ms；扫描电荷＝1-6；高能碰撞(top 20)：分辨率＝17500；隔离窗口＝2质荷比；离子捕获数量＝1e5；最大注入时间＝45ms；碰撞能量＝28,动态排除时间＝30s；
(5)分子对接筛选：通过分子对接软件AutodockVina对所得肽序进行模拟对接，筛选出4条与酪氨酸酶螯合最为紧密的肽，这4条肽的序列分别为FSGLD，PFRMY，RGFTGM和APGAPGGGF，并利用Pymol及Discovery Studio软件进行可视化处理。
图1为五肽FSGLD与酪氨酸酶的螯合图，图中五肽FSGLD与酪氨酸酶的CYS83及2个铜离子存在氢键作用力，说明五肽FSGLD进入酶活性中心与铜离子相互作用，从而抑制酶活性；图2为五肽PFRMY与酪氨酸酶的螯合图，图中五肽PFRMY与酪氨酸酶的HIS85、HIS244、ASN260、TYR65存在氢键作用力，说明五肽PFRMY与酪氨酸酶活性中心的组氨酸螯合紧密，既覆盖了催化位点，同时抑制了铜离子的催化活性；图3为六肽RGFTGM与酪氨酸酶的螯合图，图中六肽RGFTGM与酪氨酸酶的HIS85及SER282存在氢键作用力，说明六肽RGFTGM进入酶疏水空腔与铜离子附近的疏水性氨基酸螯合，占据酶与底物的作用位点；图4为九肽APGAPGGGF与酪氨酸酶的螯合图，图中九肽APGAPGGGF与酪氨酸酶的HIS244及ASN81存在氢键作用力，说明九肽APGAPGGGF进入酶疏水空腔与铜离子附近的疏水性氨基酸螯合，占据酶与底物的作用位点。
图5为固定化铜锌离子亲和层析色谱图。由图5可知，纯化罗非鱼皮肽组分的保留时间为38min。
相关测定方法介绍如下：
酪氨酸酶抑制率的测定方法：反应在96孔板中进行，反应体系为200μL。以磷酸缓冲盐溶液(PBS缓冲液)配制1mg/mL的L-多巴溶液，PBS缓冲液的pH值为6.8，浓度为0.2M。加入80μL的L-多巴溶液，再加入同等体积的酶解物(浓度为20mg/mL)，预培养5min后测定475nm处吸光值。最后加入40μL酪氨酸酶溶液(500U/mL)，37℃孵育15min再次测定475nm的吸光值。对照组以PBS缓冲液替代酶解物。酪氨酸酶抑制率按以下公式进行计算：
酪氨酸酶抑制率(％)＝(1-(A2′-A2)/(A1′-A1))×100，式中：A1，A2分别为不加酶解物组与加酶解物组预培养5min的吸光值；A1′，A2′分别为两组培养15min后的吸光值。
表1鱼皮胶原、碱性蛋白酶酶解物、纯化罗非鱼皮肽组分及纯肽的酪氨酸酶抑制率
由表1可知，罗非鱼皮胶原的酪氨酸酶抑制活性较低，其半抑制率IC50为50.13mg/mL；经过碱性蛋白酶酶解后其抑制活性显著提高，半抑制率IC50为19.87mg/mL；进一步通过固定化铜锌离子亲和层析纯化后其酪氨酸酶抑制率较鱼皮胶原提高了5倍。对液相色谱-串联质谱联用技术鉴定的肽序列进行分子对接筛选得到4条具有最高预测活性的肽，其酪氨酸酶抑制率较纯化罗非鱼皮肽组分提高了12倍以上，4条肽的酪氨酸酶抑制活性排序为PFRMY＞FSGLD＞RGFTGM＞APGAPGGGF。
实施例2
(1)取罗非鱼皮，加入0.2mol/L NaOH溶液，加入罗非鱼皮和NaOH溶液的质量比为1:10，浸泡30min后流水洗至中性；再加入0.2mol/L盐酸，加入罗非鱼皮和盐酸的质量比为1:10，浸泡30min后流水洗至中性，沥干打浆，将浆液置于65℃恒温水浴，8h后离心取上清液浓缩，得罗非鱼皮胶原提取液；
(2)取鱼皮胶原，加入胰蛋白酶，胰蛋白酶的添加量为罗非鱼皮胶原提取液中蛋白质质量的2％，在pH8.0，37℃条件下保温震荡反应4h，反应结束后沸水浴灭酶15min，抽滤，再以转速为10000r/min离心30min，取上清液干燥，得罗非鱼皮酶解粉；
(3)将罗非鱼皮酶解粉溶解在缓冲液中制成酶解液，以0.6mol/L硫酸铜溶液与0.2mol/L硫酸锌溶液等体积混合制备柱，将酶解液通过固定化铜锌离子亲和层析柱，上样量为10mg/次，螯合时间为90min，用pH值为7.6的缓冲液洗去未螯合组分，再用pH值为4.0且含NaCl浓度为0.5mol/L的溶液作为洗脱液洗脱螯合肽，洗脱速度为5mL/min，冷冻干燥，得到纯化罗非鱼皮肽组分冻干样，即为具有铜锌离子螯合活性的罗非鱼皮肽；
(4)将纯化罗非鱼皮肽组分冻干样经由配备在线纳喷离子源的液相色谱-串联质谱联用分析，液相色谱仪上样体积为10μL，样品分离梯度为40min，柱流量控制在500nL/min，流动相A相：0.1％甲酸水溶液，B相：含0.1％甲酸的乙腈溶液，柱温为40℃，电喷雾电压2kV；质谱仪参数设置如下：MS:质荷比扫描范围＝100-1500；分辨率＝70000；离子捕获数量＝3e6；最大注入时间＝50ms；扫描电荷＝1-6；高能碰撞(top20)：分辨率＝17500；隔离窗口＝2质荷比；离子捕获数量＝1e5；最大注入时间＝45ms；碰撞能量＝28,动态排除时间＝30s；
(5)分子对接筛选：通过分子对接软件AutodockVina对所得肽序进行模拟对接，筛选出4条与酪氨酸酶螯合最为紧密的肽，这4条肽的序列分别为FSGLD，PFRMY，RGFTGM和APGAPGGGF，并利用Pymol及Discovery Studio软件进行可视化处理。
实施例3
(1)取罗非鱼皮，加入0.1mol/L NaOH溶液，加入罗非鱼皮和NaOH溶液的质量比为1:8，浸泡25min后流水洗至中性；再加入0.1mol/L盐酸，加入罗非鱼皮和盐酸的质量比为1:8，浸泡20min后流水洗至中性，沥干打浆，将浆液置于60℃恒温水浴，5h后离心取上清液浓缩，得罗非鱼皮胶原提取液；
(2)取鱼皮胶原，加入木瓜蛋白酶，木瓜蛋白酶的添加量为罗非鱼皮胶原提取液中蛋白质质量的1％，在pH6.5，25℃条件下保温震荡反应8h，反应结束后沸水浴灭酶15min，抽滤，再以转速为9000r/min离心20min，取上清液干燥，得罗非鱼皮酶解粉；
(3)将罗非鱼皮酶解粉溶解在缓冲液中制成酶解液，以0.4mol/L硫酸铜溶液与0.2mol/L硫酸锌溶液等体积混合制备柱，将酶解液通过固定化铜锌离子亲和层析柱，上样量为8mg/次，螯合时间为60min，用pH值为7.6的缓冲液洗去未螯合组分，再用pH值为4.0且含NaCl浓度为0.5mol/L的溶液作为洗脱液洗脱螯合肽，洗脱速度为3mL/min，冷冻干燥，得到纯化罗非鱼皮肽组分冻干样，即为具有铜锌离子螯合活性的罗非鱼皮肽；
(4)将纯化罗非鱼皮肽组分冻干样经由配备在线纳喷离子源的液相色谱-串联质谱联用分析，液相色谱仪上样体积为8μL，样品分离梯度为30min，柱流量控制在400nL/min，流动相A相：0.1％甲酸水溶液，B相：含0.1％甲酸的乙腈溶液，柱温为40℃，电喷雾电压2kV；质谱仪参数设置如下：MS:质荷比扫描范围＝100-1500；分辨率＝70000；离子捕获数量＝3e6；最大注入时间＝50ms；扫描电荷＝1-6；高能碰撞(top20)：分辨率＝17500；隔离窗口＝2质荷比；离子捕获数量＝1e5；最大注入时间＝45ms；碰撞能量＝28，动态排除时间＝30s；
(5)分子对接筛选：通过分子对接软件AutodockVina对所得肽序进行模拟对接，筛选出4条与酪氨酸酶螯合最为紧密的肽，这4条肽的序列分别为FSGLD，PFRMY，RGFTGM和APGAPGGGF，并利用Pymol及Discovery Studio软件进行可视化处理。
实施例4
从Protein Data Bank(PDB)数据库获取酪氨酸酶的3D结构(PDB ID：2Y9X)并将其作为受体蛋白，去除受体蛋白的配体及水分子并添加氢原子。采用Chem Draw软件分别绘制五肽FSGLD、五肽PFRMY、六肽RGFTGM和九肽APGAPGGGF的二级结构并利用Minimize Energy模块进行能量最小化处理。AutoDockVina软件中设置网格地图大小为
对接中心坐标值设置为X：-10.09；Y：-28.03；Z：-43.14。以“-Docker Energy”值为筛选指标，该值越小说明酪氨酸酶抑制活性越高，得到4条最具潜在酪氨酸酶抑制活性的肽FSGLD，PFRMY，RGFTGM和APGAPGGGF。4条肽与酪氨酸酶螯合的作用机制通过Discovery Studio软件呈现可视化。
表2四条胶原蛋白源酪氨酸酶抑制肽的分子量及分子对接结果
对液相色谱-串联质谱联用技术鉴定的肽序列进行分子对接筛选得到4条具有最高活性的肽，其分子量大小与对接能如表2所示。对接能越小说明酪氨酸酶抑制活性越高。分子对接结果预测4条肽的酪氨酸酶抑制活性排序如下：PFRMY＞FSGLD＞RGFTGM＞APGAPGGGF，与实际测得的活性结果一致，说明采用分子对接方法筛选高活性酪氨酸酶抑制活性结果可靠。此外，该表亦说明小分子多肽较大分子多肽具有更高的酪氨酸酶抑制活性。
实施例5
实施例制备的胶原蛋白源酪氨酸酶抑制肽在美白面霜中的应用。
含有罗非鱼皮胶原蛋白源酪氨酸酶抑制肽的美白面霜的制备方法，由如下成分制成，按质量份数计：甘油15份，玫瑰精油10份，胶原蛋白源酪氨酸酶抑制肽10份，丁二醇5份，改性硅油5份，沙棘籽油5份，乳化剂5份，烟酰胺3份，维生素C 3份，熊果苷2份，维生素E 2份，透明质酸钠2份，增稠剂2份，吐温80 2份，山梨醇1份，苯氧乙醇0.3份，余量为去离子水。
以上实施例仅用于阐述本发明，而本发明的保护范围并非仅仅局限于以上实施例。在本发明的精神和权利要求的保护范围内，对本发明作出的任何修改和改变，都落入本发明的保护范围。
序列表
<110> 中国水产科学研究院南海水产研究所
<120> 一种胶原蛋白源酪氨酸酶抑制肽及其制备方法与应用
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 5
<212> PRT
<213> 罗非鱼(Oreochromis mossambicus)
<400> 1
Phe Ser Gly Leu Asp
1 5
<210> 2
<211> 5
<212> PRT
<213> 罗非鱼(Oreochromis mossambicus)
<400> 2
Pro Phe Arg Met Tyr
1 5
<210> 3
<211> 6
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<213> 罗非鱼(Oreochromis mossambicus)
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<212> PRT
<213> 罗非鱼(Oreochromis mossambicus)
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Ala Pro Gly Ala Pro Gly Gly Gly Phe
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Claims (2)
1.一种胶原蛋白源酪氨酸酶抑制肽，其特征在于，所述胶原蛋白源酪氨酸酶抑制肽的氨基酸序列为RGFTGM或APGAPGGGF。
2.权利要求1所述一种胶原蛋白源酪氨酸酶抑制肽在美白面霜中的应用。
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