Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02540, USA; Institut des Sciences de la Vie, Université Catholique de Louvain, Louvain-la-Neuve 1348, Belgium.
Biology Department, Woods Hole Oceanographic Institution, Woods Hole, MA 02540, USA; Boston University Superfund Research Program, Boston University School of Public Health, Boston, MA, USA.
Biochim Biophys Acta Proteins Proteom. 2018 Jan;1866(1):155-165. doi: 10.1016/j.bbapap.2017.06.026. Epub 2017 Jul 8.
Limited knowledge of the molecular evolution of deep-sea fish proteomes so far suggests that a few widespread residue substitutions in cytosolic proteins binding hydrophilic ligands contribute to resistance to the effects of high hydrostatic pressure (HP). Structure-function studies with additional protein systems, including membrane bound proteins, are essential to provide a more general picture of adaptation in these extremophiles. We explored molecular features of HP adaptation in proteins binding hydrophobic ligands, either in lipid bilayers (cytochrome P450 1A - CYP1A) or in the cytosol (the aryl hydrocarbon receptor - AHR), and their partners P450 oxidoreductase (POR) and AHR nuclear translocator (ARNT), respectively. Cloning studies identified the full-length coding sequence of AHR, CYP1A and POR, and a partial sequence of ARNT from Coryphaenoides armatus, an abyssal gadiform fish thriving down to 5000m depth. Inferred protein sequences were aligned with many non-deep-sea homologs to identify unique amino acid substitutions of possible relevance in HP adaptation. Positionally unique substitutions of various physicochemical properties were found in all four proteins, usually at sites of strong-to-absolute residue conservation. Some were in domains deemed important for protein-protein interaction or ligand binding. In addition, some involved removal or addition of beta-branched residues; local modifications of beta-branched residue patterns could be important to HP adaptation. In silico predictions further suggested that some unique substitutions might substantially modulate the flexibility of the polypeptide segment in which they are found. Repetitive motifs unique to the abyssal fish AHR were predicted to be rich in glycosylation sites, suggesting that post-translational changes could be involved in adaptation as well. Recombinant CYP1A and AHR showed functional properties (spectral characteristics, catalytic activity and ligand binding) that demonstrate proper folding at 1atm, indicating that they could be used as deep-sea fish protein models to further evaluate protein function under pressure. This article is part of a Special Issue entitled: Cytochrome P450 biodiversity and biotechnology, edited by Erika Plettner, Gianfranco Gilardi, Luet Wong, Vlada Urlacher, Jared Goldstone".
深海鱼类蛋白质组分子进化的知识有限，目前的研究表明，胞质蛋白结合亲水配体的几个广泛存在的残基取代有助于抵抗高静水压力（HP）的影响。包括膜结合蛋白在内的更多蛋白质系统的结构-功能研究对于提供这些极端微生物适应的更全面的图景至关重要。我们研究了结合疏水性配体的蛋白质（脂质双层中的细胞色素 P450 1A-CYP1A 和细胞质中的芳烃受体-AHR）适应 HP 的分子特征，以及它们各自的伴侣 P450 氧化还原酶（POR）和芳烃受体核转位蛋白（ARNT）。克隆研究从深海鳕形目鱼类矛尾鱼 Coryphaenoides armatus 中鉴定出 AHR、CYP1A 和 POR 的全长编码序列以及 ARNT 的部分序列。推断的蛋白质序列与许多非深海同源物进行了比对，以鉴定可能与 HP 适应相关的独特氨基酸取代。在所有四种蛋白质中，都发现了具有各种物理化学性质的独特取代位置，通常位于强到绝对残基保守的部位。有些位于被认为对蛋白质-蛋白质相互作用或配体结合重要的结构域中。此外，一些涉及到β-支链残基的去除或添加；β-支链残基模式的局部修饰对于 HP 适应可能很重要。计算机预测进一步表明，一些独特的取代可能会极大地调节它们所在多肽段的柔韧性。深渊鱼 AHR 特有的重复基序被预测富含糖基化位点，表明翻译后修饰也可能参与适应。重组 CYP1A 和 AHR 表现出功能特性（光谱特征、催化活性和配体结合），表明它们在 1atm 下可以正确折叠，这表明它们可以用作深海鱼类蛋白质模型，以进一步评估压力下的蛋白质功能。本文是题为“细胞色素 P450 生物多样性和生物技术”的特刊的一部分，由 Erika Plettner、Gianfranco Gilardi、Luet Wong、Vlada Urlacher 和 Jared Goldstone 编辑。
