Laboratório de Fisiologia e Bioquímica de Microrganismos, (LFBM), Universidade Estadual do Norte Fluminense Darcy Ribeiro, Campos do Goytacazes, RJ, Brazil.
Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de Sao Paulo, Sao Paulo, SP, Brazil.
Redox Biol. 2024 Jun;72:103128. doi: 10.1016/j.redox.2024.103128. Epub 2024 Mar 28.
YbbN/CnoX are proteins that display a Thioredoxin (Trx) domain linked to a tetratricopeptide domain. YbbN from Escherichia coli (EcYbbN) displays a co-chaperone (holdase) activity that is induced by HOCl. Here, we compared EcYbbN with YbbN proteins from Xylella fastidiosa (XfYbbN) and from Pseudomonas aeruginosa (PaYbbN). EcYbbN presents a redox active Cys residue at Trx domain (Cys63), 24 residues away from SQHC motif (SQHC[N]C) that can form mixed disulfides with target proteins. In contrast, XfYbbN and PaYbbN present two Cys residues in the CXXC (CAPC) motif, while only PaYbbN shows the Cys residue equivalent to Cys63 of EcYbbN. Our phylogenetic analysis revealed that most of the YbbN proteins are in the bacteria domain of life and that their members can be divided into four groups according to the conserved Cys residues. EcYbbN (SQHC[N]C), XfYbbN (CAPC[N]V) and PaYbbN (CAPC[N]C) are representatives of three sub-families. In contrast to EcYbbN, both XfYbbN and PaYbbN: (1) reduced an artificial disulfide (DTNB) and (2) supported the peroxidase activity of Peroxiredoxin Q from X. fastidiosa, suggesting that these proteins might function similarly to the canonical Trx enzymes. Indeed, XfYbbN was reduced by XfTrx reductase with a high catalytic efficiency (k/K = 1.27 x 10 M s), similar to the canonical XfTrx (XfTsnC). Furthermore, EcYbbN and XfYbbN, but not PaYbbN displayed HOCl-induced holdase activity. Remarkably, EcYbbN gained disulfide reductase activity while lost the HOCl-activated chaperone function, when the SQHC was replaced by CQHC. In contrast, the XfYbbN CAPA mutant lost the disulfide reductase activity, while kept its HOCl-induced chaperone function. In all cases, the induction of the holdase activity was accompanied by YbbN oligomerization. Finally, we showed that deletion of ybbN gene did not render in P. aeruginosa more sensitive stressful treatments. Therefore, YbbN/CnoX proteins display distinct properties, depending on the presence of the three conserved Cys residues.
YbbN/CnoX 是一类含有硫氧还蛋白（Trx）结构域和四肽重复结构域的蛋白。大肠杆菌（EcYbbN）的 YbbN 具有分子伴侣（伴侣酶）活性，这种活性由 HOCl 诱导。在这里，我们将 EcYbbN 与来自木糖氧化无色杆菌（XfYbbN）和铜绿假单胞菌（PaYbbN）的 YbbN 蛋白进行了比较。EcYbbN 的 Trx 结构域（Cys63）有一个氧化还原活性半胱氨酸残基，距离 SQHC 基序（SQHC[N]C）24 个氨基酸，可与靶蛋白形成混合二硫键。相比之下，XfYbbN 和 PaYbbN 的 CXXC（CAPC）基序中有两个半胱氨酸残基，而只有 PaYbbN 具有相当于 EcYbbN 的 Cys63 的半胱氨酸残基。我们的系统发育分析表明，大多数 YbbN 蛋白存在于细菌生命域，并且根据保守半胱氨酸残基的存在，它们的成员可以分为四个亚群。EcYbbN（SQHC[N]C）、XfYbbN（CAPC[N]V）和 PaYbbN（CAPC[N]C）是三个亚家族的代表。与 EcYbbN 不同，XfYbbN 和 PaYbbN：（1）还原了一个人工二硫键（DTNB），（2）支持木糖氧化无色杆菌的过氧化物酶 Q 的过氧化物酶活性，这表明这些蛋白可能具有与典型的 Trx 酶类似的功能。事实上，XfYbbN 被 XfTrx 还原酶以高催化效率（k/K=1.27 x 10^5 M^-1 s^-1）还原，类似于典型的 XfTrx（XfTsnC）。此外，EcYbbN 和 XfYbbN，但不是 PaYbbN，表现出 HOCl 诱导的伴侣酶活性。值得注意的是，当 SQHC 被 CQHC 取代时，EcYbbN 获得了二硫键还原酶活性，而失去了 HOCl 激活的伴侣酶功能。相比之下，XfYbbN 的 CAPA 突变体失去了二硫键还原酶活性，但保持了其 HOCl 诱导的伴侣酶功能。在所有情况下，伴侣酶活性的诱导伴随着 YbbN 寡聚化。最后，我们表明，铜绿假单胞菌中 ybbN 基因的缺失并没有使其对应激处理更敏感。因此，YbbN/CnoX 蛋白根据三个保守半胱氨酸残基的存在表现出不同的特性。
