Department of Biological and Biochemical Sciences, Faculty of Chemical Technology, University of Pardubice, 532 10 Pardubice, Czech Republic.
Gymnasium, Pardubice, Mozartova, 530 09 Pardubice, Czech Republic.
FEMS Yeast Res. 2019 Jan 1;19(1). doi: 10.1093/femsyr/foy112.
Most of the phosphatases of human fungal pathogens Candida albicans and C. parapsilosis have never been experimentally characterised, although dephosphorylation reactions are central to many biological processes. PHO15 genes of these yeasts have been annotated as the sequences encoding 4-nitrophenyl phosphatase, on the basis of homology to PHO13 gene from the bakers' yeast Saccharomyces cerevisiae. To examine the real function of these potential phosphatases from Candida spp., CaPho15p and CpPho15p were prepared using expression in Escherichia coli and characterised. They share the hallmark motifs of the haloacid dehalogenase superfamily, readily hydrolyse 4-nitrophenyl phosphate at pH 8-8.3 and require divalent cations (Mg2+, Mn2+ or Co2+) as cofactors. CaPho15p and CpPho15p did not dephosphorylate phosphopeptides, but rather hydrolysed molecules related to carbohydrate metabolism. The preferred substrate for the both phosphatases was 2-phosphoglycolate. Among the other molecules tested, CaPho15 showed preference for glyceraldehyde phosphate and ß-glycerol phosphate, while CpPho15 dephosphorylated mainly 1,3-dihydroxyacetone phosphate. This type of substrate specificity indicates that CaPho15 and CpPho15 may be a part of metabolic repair system of C. albicans and C. parapsilosis.
大多数人类真菌病原体白色念珠菌和近平滑念珠菌的磷酸酶从未经过实验鉴定，尽管去磷酸化反应是许多生物过程的核心。这些酵母的 PHO15 基因已被注释为编码 4-硝基苯磷酸酶的序列，这是基于与酿酒酵母 PHO13 基因的同源性。为了研究这些来自念珠菌属的潜在磷酸酶的实际功能，使用大肠杆菌中的表达制备了 CaPho15p 和 CpPho15p，并对其进行了表征。它们都具有卤代酸脱卤酶超家族的标志性基序，在 pH8-8.3 下容易水解 4-硝基苯磷酸盐，并需要二价阳离子（Mg2+、Mn2+或 Co2+）作为辅助因子。CaPho15p 和 CpPho15p 不能使磷酸肽去磷酸化，而是水解与碳水化合物代谢有关的分子。两种磷酸酶的首选底物是 2-磷酸甘油酸。在测试的其他分子中，CaPho15 对甘油醛磷酸盐和β-甘油磷酸表现出偏好，而 CpPho15 主要磷酸化 1,3-二羟基丙酮磷酸。这种类型的底物特异性表明 CaPho15 和 CpPho15 可能是白色念珠菌和近平滑念珠菌代谢修复系统的一部分。
