THE EUKARYOTIC HOST FACTOR THAT ACTIVATES EXOENZYME-S OF PSEUDOMONAS-AERUGINOSA IS A MEMBER OF THE 14-3-3 PROTEIN FAMILY
THE EUKARYOTIC HOST FACTOR THAT ACTIVATES EXOENZYME-S OF PSEUDOMONAS-AERUGINOSA IS A MEMBER OF THE 14-3-3 PROTEIN FAMILY
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DOI：
10.1073/pnas.90.6.2320
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发表时间：
1993-03-15
影响因子：
11.1
通讯作者：
COLLIER, RJ
中科院分区：
文献类型：
--
作者：
FU, H;COBURN, J;COLLIER, RJ
外切酶S(ExOS)是铜绿假单胞菌的一个毒力因子，它催化NAD+的腺苷二磷酸核糖部分转移到许多真核细胞蛋白上。它的首选底物包括RAS和其他几种21到25 kDa的GTP结合蛋白。EXOS绝对需要一种普遍存在的真核蛋白因子，称为Fas(激活EXOS的因子)，才能发挥酶的活性。在这里，我们描述了从牛脑cDNA文库中克隆和表达Fas的编码基因，并证明了在大肠杆菌中产生的纯化的重组Fas在一个确定的无细胞系统中激活EXOS。推导的氨基酸序列表明，该蛋白(245个残基，计算相对分子质量27,743 Da)属于高度保守、分布广泛的真核蛋白家族，统称为14-3-3蛋白。已报道的14-3-3家族成员的多种功能，包括磷脂酶A2活性和调节酪氨酸羟基酶、色氨酸羟基酶，可能还有蛋白激酶C的活性。Fas是一种14-3-3蛋白，这为该蛋白家族建立了一个额外的功能--激活外源ADP-核糖基转移酶。阐明Fas在激活EXOS中的确切作用将有助于理解铜绿假单胞菌致病的分子机制。
Exoenzyme S (ExoS), which has been implicated as a virulence factor of Pseudomonas aeruginosa, catalyzes transfer of the ADP-ribose moiety of NAD+ to many eukaryotic cellular proteins. Its preferred substrates include Ras and several other 21- to 25-kDa GTP-binding proteins. ExoS absolutely requires a ubiquitous eukaryotic protein factor, termed FAS (factor activating ExoS), for enzymatic activity. Here we describe the cloning and expression of a gene encoding FAS from a bovine brain cDNA library and demonstrate that purified recombinant FAS produced in Escherichia coli activates ExoS in a defined cell-free system. The deduced amino acid sequence of FAS shows that the protein (245 residues, calculated molecular mass 27,743 Da) belongs to a highly conserved, widely distributed eukaryotic protein family, collectively designated as 14-3-3 proteins. Various functions have been reported for members of the 14-3-3 family, including phospholipase A2 activity and regulation of tyrosine hydroxylase, tryptophan hydroxylase, and, possibly, protein kinase C activities. Identification of FAS as a 14-3-3 protein establishes an additional function for this family of proteins-the activation of an exogenous ADP-ribosyltransferase. Elucidation of the precise role of FAS in activating ExoS will contribute to understanding the molecular mechanisms by which P. aeruginosa causes disease.