Ozaki Y, Iwata J, Ohashi T
Blood. 1984 Nov;64(5):1094-102.
Wheat germ agglutinin (WGA) stimulated neutrophils to produce significant levels of luminol-dependent chemiluminescence (CL). Since WGA is known to bind N-acetylglucosamine (GlcNAc) oligomers and N-acetylneuraminic acid (NANA), we attempted to determine which binding property of WGA is essential for induction of CL. The succinylated form of WGA (SuWGA), which is no longer able to bind NANA, was still able to induce CL. N-Acetylglucosamine at a concentration of 20 mmol/L almost completely inhibited WGA-induced CL production by neutrophils, whereas bovine submaxillary gland mucin, a potent blocker of NANA binding of WGA, failed to inhibit CL production. Lectins with the GlcNAc-binding property were examined for their ability to induce CL. Those that have higher valences and have a tendency to bind GlcNAc oligomers in the internal portion of glycoconjugates were able to induce CL, whereas those that have low valences and bind terminal GlcNAc of glycoconjugates failed to induce CL even at high concentrations. Attempts were made to characterize the neutrophil membrane proteins recognized by WGA. Glycoproteins with a molecular weight of 25,000 daltons were identified by a 50 mmol/L GlcNAc elution of WGA gels loaded with 125I-labeled neutrophil membrane proteins. Elution with 500 mumol/L GlcNAc trimer produced several glycoproteins of different molecular weights in addition to the glycoproteins of 25,000 daltons. 125I-labeled WGA and SuWGA were used for autoradiographic analysis of cell extracts of the neutrophils separated on sodium dodecyl sulfate polyacrylamide gels. WGA recognized multiple glycoproteins of different molecular weights, whereas SuWGA bound only a few of them. Glycoproteins of 25,000 daltons, probably corresponding to those identified by 50 mmol/L GlcNAc elution, were also recognized.
小麦胚凝集素（WGA）刺激中性粒细胞产生显著水平的鲁米诺依赖性化学发光（CL）。由于已知WGA能结合N-乙酰葡糖胺（GlcNAc）寡聚物和N-乙酰神经氨酸（NANA），我们试图确定WGA的哪种结合特性对于诱导CL至关重要。不再能够结合NANA的琥珀酰化形式的WGA（SuWGA）仍能够诱导CL。浓度为20 mmol/L的N-乙酰葡糖胺几乎完全抑制WGA诱导的中性粒细胞CL产生，而牛颌下腺粘蛋白，一种WGA与NANA结合的有效阻断剂，未能抑制CL产生。研究了具有GlcNAc结合特性的凝集素诱导CL的能力。那些具有较高价态且倾向于结合糖缀合物内部GlcNAc寡聚物的凝集素能够诱导CL，而那些具有低价态且结合糖缀合物末端GlcNAc的凝集素即使在高浓度下也未能诱导CL。尝试对被WGA识别的中性粒细胞膜蛋白进行表征。通过用50 mmol/L GlcNAc洗脱加载有125I标记的中性粒细胞膜蛋白的WGA凝胶，鉴定出分子量为25,000道尔顿的糖蛋白。用500 μmol/L GlcNAc三聚体洗脱除了25,000道尔顿的糖蛋白外还产生了几种不同分子量的糖蛋白。125I标记的WGA和SuWGA用于对在十二烷基硫酸钠聚丙烯酰胺凝胶上分离的中性粒细胞细胞提取物进行放射自显影分析。WGA识别多种不同分子量的糖蛋白，而SuWGA仅结合其中少数几种。25,000道尔顿的糖蛋白，可能与通过50 mmol/L GlcNAc洗脱鉴定出的那些相对应，也被识别。
