Structure and function of fungal adhesins

GPI连接的细胞壁蛋白家族，称为真菌粘附素，在芽殖酵母酿酒酵母、人类致病酵母光滑念珠菌和工业相关酵母巴斯德毕赤酵母中赋予独特的表面粘附特性。迄今为止，这些赋予表面识别和粘附的蛋白质的大多数详细特征在很大程度上是未知的。在上一个资助期间，我们确定了S。酿酒酵母絮凝蛋白Flo5和Flo11以及C.光滑苔粘附素Epa1的原子分辨率。在下一个资助期，我们建议进一步阐明S. cerevisiae，C. glabrata和巴斯德毕赤酵母在体外和体内。具体来说，我们将分析从C. Glabrata Epa和Pwp家族以及来自巴斯德毕赤酵母的Flo5相关的推定粘附素。将通过聚糖微阵列分析筛选各个重组粘附素结构域的特异性配体结合，并通过X射线晶体学分析进行结晶和结构测定。不同粘附素配体结合域的体内性质将通过使用S.酿酒酵母作为异源系统。我们还将确定真菌粘附素的中心重复B结构域的生化性质，其结构和功能还不清楚，通过使用S。酿酒酵母絮凝蛋白Flo5作为模型。

A family of GPI-linked cell wall proteins, known as fungal adhesins, confers unique surface adhesion properties in the budding yeast Saccharomyces cerevisiae, the human pathogenic yeast Candida glabrata and the industrially relevant yeast Pichia pastoris. The detailed features of most of these proteins conferring surface discrimination and adherence were so far largely unknown. During the last funding period, we determined crystal structures of the adhesion domains of the S. cerevisiae floccu-lins Flo5 and Flo11 as well as of the C. glabrata adhesin Epa1 at atomic resolutions. For the next funding period, we propose to elucidate the structural and functional properties of further carbohy-drate-binding adhesins from S. cerevisiae, C. glabrata and P. pastoris in vitro and in vivo. Specifically, we will analyze N-terminal ligand-binding domains of selected members from the C. glabrata Epa and Pwp families and of Flo5-related putative adhesins from P. pastoris. Respective recombinant adhesin domains will be screened for specific ligand binding by glycan microarray analysis and subjected to crystallization and structure determination by X-ray crystallographic analysis. The in vivo properties of the different adhesin ligand-binding domains will further be investigated by functional analysis using S. cerevisiae as a heterologous system. We will also determine the biochemical properties of the central, repetitive B domains of fungal adhesins, whose structures and functions are not known in detail yet, by using the S. cerevisiae flocculin Flo5 as a model.