The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase of Candida albicans is a surface antigen
The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase of Candida albicans is a surface antigen
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DOI：
10.1128/jb.179.16.4992-4999.1997
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发表时间：
1997-08-01
影响因子：
3.2
通讯作者：
Gozalbo, D
中科院分区：
文献类型：
--
作者：
GilNavarro, I;Gil, ML;Gozalbo, D
从白色念珠菌ATCC 26555的λ gt 11 cDNA文库中筛选了两名系统性念珠菌病患者和五名具有高水平抗-C的血小板减少症患者的混合血清。从60，000个重组质粒中分离出7个克隆。最具反应性的一个含有0.9kb的cDNA，其编码仅与系统性念珠菌病患者血清免疫反应的多肽。编码区的核苷酸序列与酿酒酵母（Saccharomycescerevisiae）编码甘油醛-3-磷酸脱氢酶（GAPDH）的基因TDH 1至TDH 3具有78 - 79%的同源性，氨基酸序列具有76%的同源性，因此将该基因命名为C.白色念珠菌TDH 1.制备了兔抗胞质C.用抗CA-GAPDH的多克隆抗体（PAb）鉴定含细胞壁部分的β-巯基乙醇提取物中的GAPDH。白色念珠菌细胞表面，特别是芽生孢子上。半定量流式细胞术分析表明，这种GAPDH形式的敏感性胰蛋白酶和其电阻被删除与2 M NaCl或2%十二烷基硫酸钠，荧光的减少在可溶性GAPDH的存在下表明标记的特异性，此外，剂量依赖性GAPDH酶活性检测在完整的芽生孢子和芽管细胞。用胰蛋白酶、甲醛和PAb抗CA-GAPDH预处理细胞可降低该活性。这些观察结果表明，免疫原性，酶活性细胞壁相关形式的糖酵解酶GAPDH被发现在细胞表面的C。白色念珠菌细胞
A lambda gt11 cDNA library from Candida albicans ATCC 26555 Has screened by using pooled sera from two patients with systemic candidiasis and five neutropenic patients with high levels of anti-C. albicans immunoglobulin M antibodies, Seven clones were isolated from 60,000 recombinant phages. The most reactive one contained a 0.9-kb cDNA encoding a polypeptide immunoreactive only with sera from patients with systemic candidiasis, The whole gene was isolated from a genomic library by using the cDNA as a probe. The nucleotide sequence of the coding region showed homology (78 to 79%) to the Saccharomyces cerevisiae TDH1 to TDH3 genes coding for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and their amino acid sequences showed 76% identity: thus, this gene has been named C. albicans TDH1. A rabbit polyclonal antiserum against the purified cytosolic C. albicans GAPDH (polyclonal antibody [PAb] anti-CA-GAPDH) was used to identify the GAPDH in the beta-mercaptoethanol extracts containing cell wall moieties, Indirect immunofluorescence demonstrated the presence of GAPDH at the C. albicans cell surface, particularly on the blastoconidia. Semiquantitative flow cytometry analysis showed the sensitivity of this GAPDH form to trypsin and its resistance to be removed with 2 M NaCl or 2% sodium dodecyl sulfate, The decrease in fluorescence in the presence of soluble GAPDH indicates the specificity of the labelling, In addition, a dose-dependent GAPDH enzymatic activity was detected in intact blastoconidia and germ tube cells. This activity was reduced by pretreatment of the cells with trypsin, formaldehyde, and PAb anti-CA-GAPDH. These observations indicate that an immunogenic, enzymatically active cell wall-associated form of the glycolytic tic enzyme GAPDH is found at the cell surface face of C. albicans cells.