Kim Seonghun
Jeonbuk Branch Institute, Korea Research Institute of Bioscience and Biotechnology, 181 Ipsin-gil, Jeongeup 56212, Republic of Korea; Department of Biosystems and Bioengineering, KRIBB School of Biotechnology, University of Science and Technology (UST), 217 Gajeong-ro, Daejeon 34113, Republic of Korea.
Int J Biol Macromol. 2023 Jan 31;226:1010-1020. doi: 10.1016/j.ijbiomac.2022.12.091. Epub 2022 Dec 13.
The mushroom Hericium erinaceus expresses isolectins with different glycan binding specificities; of these, the ricin B-like lectin HEL1 and HEL2 (HEL2a and HEL2b) can bind fucosylated N-glycans and core 1 O-glycans, respectively. However, other lectin-like protein-coding transcripts detected in the H. erinaceus transcriptome, named HEL3, remain to be characterized. Therefore, in this study, the expression levels of all these isolectins genes were compared to characterize the molecular and biochemical properties of these carbohydrate-binding proteins. Low expression genes encoding putative cytolysin proteins, HEL3a and HEL3b, were identified. Bioinformatics analyses revealed that these proteins shared highly homologous structures and carbohydrate-binding residues with other mushroom lectins. Further, their recombinant proteins, rHEL3a and rHEL3b showed an octamer composed of identical 17 kDa subunits under non-denaturing conditions and a slightly basic isoelectric point value of approximately 8.3. The hemagglutination activity of these isolectins was strongly inhibited by glycoproteins rather than free glycans. Interestingly, glycan-binding profiles showed that rHEL3 isolectins interacted with most polylactosamine (poly-LacNAc)-extended N-glycans with relatively low binding activity. Isothermal titration calorimetry also revealed that these recombinant lectins have different binding capacities toward N-glycan-containing glycoproteins. Further, treatment with different concentrations of rHEL3 lectins showed cytotoxic effects in K562, UACC62, and CHO model cell lines, which express poly-LacNAc glycans, confirmed by inhibition of proliferation. Overall, these biochemical properties indicate that rHEL3 isolectins may be used as unique lectins for detecting poly-LacNAc-extended glycans, which are known to be over-expressed in leukemia or metastatic melanoma cells, in cancer diagnostic assays and anti-cancer therapies.
猴头菇表达具有不同聚糖结合特异性的异凝集素；其中，蓖麻毒蛋白B样凝集素HEL1和HEL2（HEL2a和HEL2b）分别可结合岩藻糖基化的N-聚糖和核心1 O-聚糖。然而，在猴头菇转录组中检测到的其他类凝集素蛋白编码转录本，命名为HEL3，其特性仍有待确定。因此，在本研究中，对所有这些异凝集素基因的表达水平进行了比较，以表征这些碳水化合物结合蛋白的分子和生化特性。鉴定出了编码假定细胞溶素蛋白的低表达基因HEL3a和HEL3b。生物信息学分析表明，这些蛋白与其他蘑菇凝集素具有高度同源的结构和碳水化合物结合残基。此外，它们的重组蛋白rHEL3a和rHEL3b在非变性条件下显示为由相同的17 kDa亚基组成的八聚体，且等电点值略呈碱性，约为8.3。这些异凝集素的血凝活性受到糖蛋白而非游离聚糖的强烈抑制。有趣的是，聚糖结合谱显示rHEL3异凝集素与大多数多乳糖胺（poly-LacNAc）延伸的N-聚糖相互作用，结合活性相对较低。等温滴定量热法还表明，这些重组凝集素对含N-聚糖的糖蛋白具有不同的结合能力。此外，用不同浓度的rHEL3凝集素处理在表达poly-LacNAc聚糖的K562、UACC62和CHO模型细胞系中显示出细胞毒性作用，这通过增殖抑制得到证实。总体而言，这些生化特性表明rHEL3异凝集素可作为独特的凝集素，用于在癌症诊断分析和抗癌治疗中检测已知在白血病或转移性黑色素瘤细胞中过度表达的poly-LacNAc延伸聚糖。
