?肖鶩軒 王昆侖 周海燕
摘 要:為了明確莼菜中鋅蛋白的分布及氨基酸組成,以富硒高鋅土壤中種植的莼菜為材料,對(duì)其幼葉蛋白質(zhì)進(jìn)行分離提取和分析。結(jié)果表明:莼菜中蛋白質(zhì)主要類型為弱堿性蛋白質(zhì),比例為44.79%,但其鋅含量相對(duì)較低,水溶性蛋白比例只有15.43%,但其鋅含量卻是弱堿性蛋白質(zhì)中的2.17倍,表明大部分鋅離子是與水溶性蛋白結(jié)合的;通過逐級(jí)鹽析從水溶性蛋白中獲得5個(gè)餾分,進(jìn)一步分析各餾分的蛋白質(zhì)含量及鋅含量,獲得WSⅡ和WSⅤ這2個(gè)高鋅含量的蛋白質(zhì)餾分,電泳分析顯示這2個(gè)餾分分別由一個(gè)或一系列相同亞基組成;氨基酸組分分析顯示,WSⅡ和WSⅤ中甘氨酸、硒代半胱氨酸、組氨酸和賴氨酸含量較高。
關(guān)鍵詞:莼菜;鋅蛋白;分離;氨基酸組成
中圖分類號(hào):Q513 文獻(xiàn)標(biāo)識(shí)碼:A 文章編號(hào):1006-060X(2020)04-0064-05
Preliminary Analysis and Separation of Zinc Protein in Brasenia schreberi
XIAO Wu-xuan1,WANG Kun-lun2,ZHOU Hai-yan2
(1. Hunan Institute of Nuclear Agriculture Science and Space Breeding, Hunan Academy of Agricultural Sciences, Changsha 410125, PRC;
2. College of Bioscience and Biotechnology, Hunan Agricultural University, Changsha 410128, PRC)
Abstract: Brasenia schreberi planted in selenium-rich and high-zinc soil was used as the target material in this study. The proteins in Brasenia schreberi young leaves were separated and analyzed for further clarification of zinc protein distribution and amino acid composition in the genus. The results show that the main type of proteins in Brasenia schreberi is weak alkaline protein, accounting for 44.79%, but its zinc content is relatively low; the proportion of water-soluble protein is only 15.43%, but its zinc content is 2.17 times that of weak alkaline protein, which indicates that most of the zinc ions are combined with water-soluble protein. Five fractions from water-soluble protein were obtained by stepwise salting out, then the protein content and zinc content of each fraction were analyzed, finally two protein fractions of high zinc content WSⅡ and WSⅤ were obtained. Electrophoresis analysis shows that these two fractions are composed of one or a series of the same subunits. Amino acid composition analysis shows that the contents of glycine, selenocysteine, histidine and lysine are high in WSⅡ and WSⅤ.
Key words: Brasenia schreberi; zinc protein; separation; amino acid composition
莼菜(Brasenia schreberi)又名蒪菜、馬蹄菜、湖菜等,系睡蓮目睡蓮科莼屬植物。莼菜為須根系植物,不定根簇生于地下匍匐莖的莖節(jié)靠近葉柄基部的兩側(cè);莖可分為3類,分別是匍匐莖、縮短莖和水中莖[1];幼葉葉片呈現(xiàn)卷曲狀態(tài),且葉外有透明質(zhì)膠包裹,可食用[2];花由花瓣、花托、花萼、花梗及雌雄蕊組成,花期為2 d,第一天呈現(xiàn)雌蕊的功能,第二天則是展現(xiàn)雄花的功能,白天花朵浮出水面,主要靠風(fēng)媒傳播授粉,夜晚花體沉入水中,于水中授粉結(jié)果,2~6周種子即可成熟[3]。莼菜在亞洲廣泛栽培,可有性繁殖,也可無性繁殖。
莼菜的營(yíng)養(yǎng)價(jià)值較高,富含可溶性植物多糖、蛋白質(zhì)及一些微量元素。目前已經(jīng)檢測(cè)到的微量元素多達(dá)27種,其中鋅元素的含量遠(yuǎn)超一般蔬菜[4]。此外,莼菜還含有膳食纖維及維生素B等物質(zhì)[4],是一種應(yīng)用前景廣闊的蔬菜。還有研究表明,莼菜提取物具有抗藻、抗菌和化感作用[5]。研究表明,莼菜多糖對(duì)小鼠的免疫系統(tǒng)具有明顯的改善作用[6-9]。
鋅是植物[10]、動(dòng)物[11]和微生物[12]必不可少的微量元素,存在于數(shù)百種特定的酶中[13]。鋅通常在轉(zhuǎn)錄因子中充當(dāng)結(jié)構(gòu)離子[14],并且通過金屬硫蛋白進(jìn)行存儲(chǔ)和轉(zhuǎn)移[15]。對(duì)人類而言,鋅有著更為普遍的生物學(xué)功能[16]。它與多種有機(jī)配體相互作用[16],并在RNA和DNA的代謝、信號(hào)轉(zhuǎn)導(dǎo)和基因表達(dá)中起作用,它還能調(diào)節(jié)細(xì)胞凋亡。除數(shù)百種參與交換和運(yùn)輸鋅的蛋白質(zhì)外,還約有10%的人類蛋白質(zhì)可與鋅結(jié)合。大量文獻(xiàn)對(duì)莼菜葉片表層凝膠和多糖的結(jié)構(gòu)和功能進(jìn)行了闡述,但對(duì)莼菜中鋅和蛋白質(zhì)的研究較少。前期研究表明,鋅在莼菜幼葉中的含量最高,并且與蛋白質(zhì)結(jié)合的鋅含量高于與多糖或其他有機(jī)分子結(jié)合的鋅含量。因此,筆者以產(chǎn)自湖北省利川市(其耕地土壤富含鋅和硒元素)的莼菜為材料,對(duì)蛋白質(zhì)進(jìn)行分離提取和分析,以期進(jìn)一步明確莼菜中鋅蛋白的含量、結(jié)構(gòu)和功能。
根據(jù)參考文獻(xiàn),用C18反相柱(250 mm×4.6 mm×5 μm)分析了WSⅡ和WSⅤ中回收的富鋅蛋白質(zhì)的氨基酸組成,結(jié)果如表1和圖5所示。從WSⅡ中回收的富鋅蛋白質(zhì)由天冬氨酸(D)、甘氨酸(G)、組氨酸(H)、精氨酸(R)、脯氨酸(P)、酪氨酸(Y)、
纈氨酸(V)、半胱氨酸(C)、硒代半胱氨酸(Se-C)、賴氨酸(K)等組成;其中,甘氨酸、酪氨酸和賴氨酸的比例較高,占比為84.98%。此外,趨于與鋅結(jié)合的組氨酸、半胱氨酸、硒代半胱氨酸、精氨酸和賴氨酸的比例也高達(dá)43.58%。從WSⅤ中回收的富鋅蛋白質(zhì)由天冬氨酸(D)、甘氨酸(G)、組氨酸(H)、蘇氨酸(T)、丙氨酸(A)、脯氨酸(P)、酪氨酸(Y)、硒代半胱氨酸(Se-C)等組成;其中,甘氨酸、酪氨酸、組氨酸、丙氨酸和硒代半胱氨酸的比例較高,占比為95.79%,而傾向于與鋅結(jié)合的組氨酸和硒代半胱氨酸的比例為12.10%。
3 討 論
該研究所選莼菜材料種植在富含鋅元素的土壤中,因此其鋅含量較一般地區(qū)高,是一種理想的鋅補(bǔ)充劑。試驗(yàn)結(jié)果表明,在該材料中,幼葉的鋅含量很高,且其傾向于與水溶性蛋白質(zhì)結(jié)合。純化后發(fā)現(xiàn),莼菜幼葉中2種鋅結(jié)合水溶性蛋白(WSⅡ和WSⅤ)分別由一個(gè)或一系列相同亞基組成。通過RT-HPLC的氨基酸分析,蛋白質(zhì)WSⅡ和WSⅤ的甘氨酸、硒代半胱氨酸和組氨酸的含量高,另外WSⅡ中的賴氨酸含量也較高。在蛋白質(zhì)中,鋅離子通常與天冬氨酸、谷氨酸、半胱氨酸和組氨酸的氨基酸側(cè)鏈配位[20]。因此,WSⅡ和WSⅤ的硒代半胱氨酸和組氨酸含量很高也就不足為奇了。硒代半胱氨酸替代半胱氨酸的原因是莼菜生長(zhǎng)的土壤中硒元素含量較高。這些結(jié)果表明,生長(zhǎng)在湖北利川地區(qū)的莼菜由于其高鋅含量及豐富的氨基酸組成而具有良好的營(yíng)養(yǎng)品質(zhì),具有較好的開發(fā)
前景。
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(責(zé)任編輯:成 平)