玉皇菇植物凝集素分離純化與特性之探討

Studies on Isolation, Purification and Characterization of Pleurotus citrinopiltus Lectin

林永順(Yeong-Shenn Lin) ； 江啟銘(Chii-Ming Jiang) ； 楊浚岳(Jiung-Yueh Yang) ； 徐嘉鴻(Chia-Hung Hsu) ； 吳明昌(Ming-Chang Wu)

《台灣農學會報》 9卷4期 (2008/08) Pp. 293-304

https://doi.org/10.6730/JAAT.200808_9(4).0001

摘要

凝集素(Lectins, Agglutinins, Affinitins, Phytohemagglutinins, Phasins Protectins)是一種結合碳水化合物之蛋白質，與醣蛋白、醣脂質或多醣有很高的親和力，具有使紅血球、正常或變形細胞發生凝集的特性。本實驗進行對玉皇菇植物凝集素分離純化並探討其一般特性，將玉皇菇子實體製成丙酮粉末(Acetone podwer)，以10%氯化鈉溶液在50℃水浴下萃取一小時，經硫酸銨沉澱劃分收集40~80%飽和度者進行DEAE-Sepharose離子交換層析，再以Sephacryl S-200膠濾層析純化，即可得到均質的玉皇搪凝集素。所得均質後萃取液用來檢測經或未經酵素(Protease, trysin, papain)處理的人類正常A、B、O型紅血球及兔子紅血球凝集反應，結果均能產生凝集現象，顯示其對人類血型紅血球並沒有特異性，蛋白酵素或胰蛋白酵素處理人類及免子紅血球，能增加凝集活性力價。當玉皇菇植物凝集素萃取時加入二價陽離子(Fe(上标 2+), Ca(上标 2+), Mg(上标 2+), Zn(上标 2+))後，其凝集活性有明顯提升的現象。以常見之17種醣與醣蛋白作醣類抑制試驗，玉皇菇值物凝集素並沒有特異醣存在。萃取液在4℃與-20℃下貯存70天，其凝集活性下降至原有活性的20%以下。此凝集素呈現活性的酸鹼值範圍廣，且具耐熱之特徵。

關鍵字

玉皇菇 ； 凝集素 ； 醣蛋白 ； 醣脂質 ； 紅血球

並列摘要

Lectins (agglutinins, affinitins, phytoagglutinins, phasins protectins) are carbohydrate proteins that can bind glycans of glycoproteins, glycolipids, or polysaccharides with high affinity, and can bind the erythrocytes, normal or transformed cell to cause agglutination. This report studied isolation, purification and characterization of Pleurotus citrinopileatus lectin. The Pleurotus citrinopileatus fruit body was homogenized with acetone, then extracted with 10% NaCl solution at 50℃. The crude lectin extract was fractionated at 40~80% saturation of ammonium sulfate then separated by DEAE-Sepharose anion exchange chromatography, and further separated with Sephacryl S-200 gel permeation chromatography. The extract was examined through agglutinin reaction with human normal A, B, and O type erythrocytes and rabbit erythrocytes. The results showed that there was no specificity on human erythrocytes treated or untreated with protease, trypsin, papain. The human and rabbit erythrocytes treated with protease and trypsin, however, could enhance the agglutination activity. The Pleurotus citrinopileatus lectin extracted with bivalent cations (Fe(superscript 2+) Ca(superscript 2+), Mg(superscript 2+), Mn(superscript 2+), Zn(superscript 2+)) could increase the agglutination activity. There was no effect of some sugars on the agglutination activity of Pleurotus citrinopileatus lectin. The agglutination activity of Pleurotus citrinopileatus lectin was lower than 20% when stored at 4 ℃ and-20 ℃ for 70 days. The lectin demonstrated that the agglutination can be contained at a wide range of pH values and temperature.

並列關鍵字

Pleurotus citrinopileatus ； Lectin ； Glycoprotein ； Glycolipid ； Erythrocytes