キムラ　マサヒロ   木村　将大    所属   応用生物学部 応用生物学科    職種   助教
言語種別	英語
発行・発表の年月	2020/04
形態種別	学術論文
査読	査読あり
標題	Development of a novel β-1,6-glucan-specific detection system using functionally modified recombinant endo-β-1,6-glucanase.
執筆形態	共著
掲載誌名	J Biol Chem
掲載区分	国外
出版社・発行元	Elsevier / American Society for Biochemistry and Molecular Biology
巻・号・頁	295(16),pp.5362-5376
著者・共著者	Yamanaka, D., Takatsu, K., Kimura, M., Swamydas, M., Ohnishi, H., Umeyama, T., Oyama, F., Lionakis MS., and Ohno, N.
概要	β-1,3-d-Glucan is a ubiquitous glucose polymer produced by plants, bacteria, and most fungi. It has been used as a diagnostic tool in patients with invasive mycoses via a highly-sensitive reagent consisting of the blood coagulation system of horseshoe crab. However, no method is currently available for measuring β-1,6-glucan, another primary β-glucan structure of fungal polysaccharides. Herein, we describe the development of an economical and highly-sensitive and specific assay for β-1,6-glucan using a modified recombinant endo-β-1,6-glucanase having diminished glucan hydrolase activity. The purified β-1,6-glucanase derivative bound to the β-1,6-glucan pustulan with a KD of 16.4 nm We validated the specificity of this β-1,6-glucan probe by demonstrating its ability to detect cell wall β-1,6-glucan from both yeast and hyphal forms of the opportunistic fungal pathogen Candida albicans, without any detectable binding to glucan lacking the long β-1,6-glucan branch. We developed a sandwich ELISA-like assay with a low limit of quantification for pustulan (1.5 pg/ml), and we successfully employed this assay in the quantification of extracellular β-1,6-glucan released by >250 patient-derived strains of different Candida species (including Candida auris) in culture supernatant in vitro We also used this assay to measure β-1,6-glucan in vivo in the serum and in several organs in a mouse model of systemic candidiasis. Our work describes a reliable method for β-1,6-glucan detection, which may prove useful for the diagnosis of invasive fungal infections.
外部リンクURL	https://www.sciencedirect.com/science/article/pii/S0021925817485553?via%3Dihub