cDNA cloning and in vitro synthesis of the Dolichos biflorus seed lectin.
cDNA cloning and in vitro synthesis of the Dolichos biflorus seed lectin.
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双花豆种子凝集素的 cDNA 克隆和体外合成。
DOI：
10.1111/j.1432-1033.1987.tb13327.x
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Etzler,ME
中科院分区：
文献类型：
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作者：
Schnell,DJ;Alexander,DC;Williams,BG;Etzler,ME
扁豆凝集素含有两个结构上相关的亚基。用提取的RNA构建了一个cDNA文库。两花种子能主动合成种子凝集素。利用lambda Charon 16载体在大肠杆菌中表达该文库，并用凝集素特异性抗血清分离种子凝集素基因。杂交种。从活性产生两个凝集素亚基的种子中分离得到的双花籽凝集素cDNAto RNA鉴定为1100个碱基的单一大小的RNA。由两个凝集素亚基共有的氨基酸序列构建的寡脱氧核苷酸探针检测到相同大小的RNA。种子RNA的体外翻译和翻译产物的免疫沉淀使用凝集素特异性抗血清产生的单一多肽的相对分子质量略高于最大的种子凝集素亚基。该种子凝集素前体与由种子凝集素cDNA选择的mRNA杂交物合成的多肽没有区别。这些数据支持THED的两种亚基类型都存在单一的多肽前体。并提出亚基类型之间的差异是翻译后加工引起的。
TheDolichos biflorusseed lectin contains two structurally related subunits. A cDNA library was constructed using RNA isolated fromD. biflorusseeds actively synthesizing the seed lectin. The library was expressed inEscherichia coliusing a lambda Charon 16 vector, and lectin‐specific antiserum was used to isolate a seed lectin cDNA. Hybridization of theD. biflorusseed lectin cDNA to RNA isolated from seeds actively producing both lectin subunits identifies a single‐size RNA of 1100 bases. An oligodeoxyribonucleotide probe, constructed from an amino acid sequence common to both lectin subunits, detects the same size RNA. Translation of seed mRNAin vitroand immunoprecipitation of translation products using a lectin‐specific antiserum yields a single polypeptide of slightly higher molecular mass than the largest seed lectin subunit. This seed lectin precursor is indistinguishable from a polypeptide synthesized from mRNA hybrid selected by the seed lectin cDNA. These data support the existence of a single polypeptide precursor for both subunit types of theD. biflorusseed lectin and suggest that differences between the subunit types arise by posttranslational processing.