Academy of Scientific and Innovative Research (AcSIR), Ghaziabad, 201002, India; CSIR-Indian Institute of Chemical Technology, Department of Applied Biology, Hyderabad, Telangana, 500007, India.
CSIR-Indian Institute of Chemical Technology, Department of Applied Biology, Hyderabad, Telangana, 500007, India.
Eur J Med Chem. 2021 Jan 1;209:112883. doi: 10.1016/j.ejmech.2020.112883. Epub 2020 Sep 28.
Methionine aminopeptidases (MetAPs) have been recognized as drug targets and have been extensively studied for discovery of selective inhibitors. MetAPs are essential enzymes in all living cells. While most prokaryotes contain a single gene, some prokaryotes and all eukaryotes including human have redundancy. Due to the similarity in the active sites of the MetAP enzyme between the pathogens and human limited the success of discovering selective inhibitors. We recently have discovered that MetAPs with small inserts within the catalytic domain to have different susceptibilities against some inhibitors compared to those that do not have. Using this clue we used bioinformatic tools to identify new variants of MetAPs with inserts in pathogenic species. Two new isoforms were identified in Vibrio species with two and three inserts in addition to an isoform without any insert. Multiple sequence alignment suggested that inserts are conserved in several of the Vibrio species. Two of the three inserts are common between two and three insert isoforms. One of the inserts is identified to have "NNKNN" motif that is similar to well-characterized quorum sensing peptide, "NNWNN". Another insert is predicted to have a posttranslational modification site. Three Vibrio proteins were cloned, expressed, purified, enzyme kinetics established and inhibitor screening has been performed. Several of the pyridinylpyrimidine derivatives selectively inhibited MetAPs with inserts compared to those that do not have, including the human enzyme. Crystal structure and molecular modeling studies provide the molecular basis for selective inhibition.
甲硫氨酸氨肽酶（MetAPs）已被认为是药物靶点，并已广泛研究用于发现选择性抑制剂。MetAPs 是所有活细胞中必不可少的酶。虽然大多数原核生物只含有一个基因，但一些原核生物和所有真核生物（包括人类）都存在冗余。由于病原体和人类的 MetAP 酶的活性位点相似，这限制了发现选择性抑制剂的成功。我们最近发现，与没有插入的 MetAP 相比，催化结构域内有小插入的 MetAP 对某些抑制剂的敏感性不同。利用这一线索，我们使用生物信息学工具在病原体物种中鉴定具有插入片段的新的 MetAP 变体。在弧菌属中鉴定出两种新的同工酶，除了没有插入片段的同工酶外，还有两种和三种插入片段。多重序列比对表明，插入片段在几种弧菌属中是保守的。三种插入片段中的两种在两种和三种插入片段同工酶之间是共同的。其中一个插入片段被鉴定为具有与特征明确的群体感应肽“NNWNN”相似的“NNKNN”基序。另一个插入片段预测具有翻译后修饰位点。克隆、表达、纯化了三种弧菌蛋白，建立了酶动力学，并进行了抑制剂筛选。与没有插入片段的酶相比，包括人类酶在内的几种吡啶并嘧啶衍生物选择性地抑制了具有插入片段的 MetAPs。晶体结构和分子建模研究为选择性抑制提供了分子基础。
