Maia Lorhenn Bryanda Lemes, Pereira Humberto D'Muniz, Garratt Richard Charles, Brandão-Neto José, Henrique-Silva Flavio, Toyama Danyelle, Dias Renata O, Bachega José Fernando Ruggiero, Peixoto Julia Vasconcellos, Silva-Filho Marcio C
Programa de Pós-Graduação em Biotecnologia, Universidade Católica Dom Bosco, Campo Grande, Brazil.
Instituto de Física de São Carlos, Universidade de São Paulo, São Carlos, Brazil.
Front Plant Sci. 2021 Sep 9;12:734248. doi: 10.3389/fpls.2021.734248. eCollection 2021.
SUGARWINs are PR-4 proteins associated with sugarcane defense against phytopathogens. Their expression is induced in response to damage by larvae. These proteins play an important role in plant defense, in particular against fungal pathogens, such as (Went) and . The pathogenesis-related protein-4 (PR-4) family is a group of proteins equipped with a BARWIN domain, which may be associated with a chitin-binding domain also known as the hevein-like domain. Several PR-4 proteins exhibit both chitinase and RNase activity, with the latter being associated with the presence of two histidine residues H11 and H113 (BARWIN) [H44 and H146, SUGARWINs] in the BARWIN-like domain. In sugarcane, similar to other PR-4 proteins, SUGARWIN1 exhibits ribonuclease, chitosanase and chitinase activities, whereas SUGARWIN2 only exhibits chitosanase activity. In order to decipher the structural determinants involved in this diverse range of enzyme specificities, we determined the 3-D structure of SUGARWIN2, at 1.55Å by X-ray diffraction. This is the first structure of a PR-4 protein where the first histidine has been replaced by asparagine and was subsequently used to build a homology model for SUGARWIN1. Molecular dynamics simulations of both proteins revealed the presence of a flexible loop only in SUGARWIN1 and we postulate that this, together with the presence of the catalytic histidine at position 42, renders it competent as a ribonuclease. The more electropositive surface potential of SUGARWIN1 would also be expected to favor complex formation with RNA. A phylogenetic analysis of PR-4 proteins obtained from 106 Embryophyta genomes showed that both catalytic histidines are widespread among them with few replacements in these amino acid positions during the gene family evolutionary history. We observe that the H11 replacement by N11 is also present in two other sugarcane PR-4 proteins: SUGARWIN3 and SUGARWIN4. We propose that RNase activity was present in the first Embryophyta PR-4 proteins but was recently lost in members of this family during the course of evolution.
甘蔗病程相关蛋白（SUGARWIN）是与甘蔗抵御植物病原体相关的病程相关蛋白4（PR-4）。它们的表达是对幼虫损伤的响应而被诱导的。这些蛋白在植物防御中发挥重要作用，特别是针对真菌病原体，如（温特）和。病程相关蛋白4（PR-4）家族是一组具有BARWIN结构域的蛋白，该结构域可能与一个也被称为类橡胶素结构域的几丁质结合结构域相关。几种PR-4蛋白同时具有几丁质酶和核糖核酸酶活性，后者与BARWIN样结构域中两个组氨酸残基H11和H113（BARWIN）[H44和H146，甘蔗病程相关蛋白（SUGARWIN）]的存在有关。在甘蔗中，与其他PR-4蛋白类似，甘蔗病程相关蛋白1（SUGARWIN1）具有核糖核酸酶、壳聚糖酶和几丁质酶活性，而甘蔗病程相关蛋白2（SUGARWIN2）仅具有壳聚糖酶活性。为了解析参与这种多种酶特异性的结构决定因素，我们通过X射线衍射在1.55埃的分辨率下确定了甘蔗病程相关蛋白2（SUGARWIN2）的三维结构。这是第一个第一个组氨酸被天冬酰胺取代的PR-4蛋白的结构，随后被用于构建甘蔗病程相关蛋白1（SUGARWIN1）的同源模型。对这两种蛋白的分子动力学模拟显示，只有甘蔗病程相关蛋白1（SUGARWIN1）存在一个柔性环，我们推测，这与第42位催化组氨酸的存在一起，使其具备作为核糖核酸酶的能力。甘蔗病程相关蛋白1（SUGARWIN1）更正电的表面电势也有望有利于与RNA形成复合物。对从106种胚植物基因组中获得的PR-4蛋白进行的系统发育分析表明，在基因家族进化历史中，这两个催化组氨酸在它们中广泛存在，在这些氨基酸位置上几乎没有替换。我们观察到，在另外两种甘蔗PR-4蛋白：甘蔗病程相关蛋白3（SUGARWIN3）和甘蔗病程相关蛋白4（SUGARWIN4）中也存在H11被N11替换的情况。我们提出，核糖核酸酶活性存在于第一个胚植物PR-4蛋白中，但在该家族成员的进化过程中最近丧失了。
