Institute for Glycomics, Griffith University, Gold Coast, QLD 4222, Australia.
Research Centre for Infectious Diseases, Department of Molecular and Biomedical Science, University of Adelaide, S.A., 5005, Australia.
Biochem Biophys Res Commun. 2018 Jun 7;500(3):765-771. doi: 10.1016/j.bbrc.2018.04.151. Epub 2018 Apr 26.
The B subunit of the subtilase cytotoxin (SubB) recognises N-glycolylneuraminic acid (Neu5Gc) containing glycans, the most prominent form of aberrant glycosylation in human cancers. We have previously engineered SubB by construction of a SubB mutant (SubB2M) for greater specificity and enhanced recognition of Neu5Gc containing glycans. In this study, we further explore the utility of SubB2M as a Neu5Gc lectin by showing its improved specificity and recognition for Neu5Gc containing glycans over the wild-type SubB protein and an anti-Neu5Gc IgY antibody in a N-acetylneuraminic acid (Neu5Ac)/Neu5Gc glycan array and by surface plasmon resonance. Far-western blot analysis showed that SubB2M preferentially binds to bovine serum glycoproteins over human serum glycoproteins. SubB2M was also able to detect Neu5Gc containing bovine glycoproteins spiked into normal human serum with greater sensitivity than the wild-type SubB and the anti-Neu5Gc IgY antibody. These results suggest that SubB2M will be a useful tool for the testing of serum and other bodily fluids for cancer diagnosis and prognosis.
枯草溶菌素 B 亚单位（SubB）的 B 亚基识别含有 N-羟乙酰神经氨酸（Neu5Gc）的聚糖，这是人类癌症中最显著的异常糖基化形式。我们之前通过构建枯草溶菌素 B 突变体（SubB2M）来提高特异性和增强对含有 Neu5Gc 的聚糖的识别，从而对 SubB 进行了工程改造。在这项研究中，我们通过在 N-乙酰神经氨酸（Neu5Ac）/Neu5Gc 聚糖阵列和表面等离子体共振中显示 SubB2M 对含有 Neu5Gc 的聚糖的特异性和识别优于野生型 SubB 蛋白和抗 Neu5Gc IgY 抗体，进一步探讨了 SubB2M 作为 Neu5Gc 凝集素的用途。远 Western blot 分析表明，SubB2M 优先结合牛血清糖蛋白而不是人血清糖蛋白。SubB2M 还能够检测到掺入正常人血清中的含有 Neu5Gc 的牛糖蛋白，其灵敏度高于野生型 SubB 和抗 Neu5Gc IgY 抗体。这些结果表明，SubB2M 将成为用于测试血清和其他体液以进行癌症诊断和预后的有用工具。
