Protein splicing of inteins with atypical glutamine and aspartate c-terminal residues
Protein splicing of inteins with atypical glutamine and aspartate c-terminal residues
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DOI：
10.1074/jbc.m311343200
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发表时间：
2004-01-30
影响因子：
4.8
通讯作者：
Pietrokovski, S
中科院分区：
文献类型：
--
作者：
Amitai, G;Dassa, B;Pietrokovski, S
内含肽是存在于许多蛋白质中的蛋白质剪接结构域。它们自我催化从宿主蛋白质中切除，通过肽键连接它们以前的侧翼。内含肽的C-末端残基通常是天冬酰胺（Asn）。这个残基环化为琥珀酰亚胺导致内含肽最终从其宿主上脱离。我们研究了两个内含肽与非典型的C-末端残基的蛋白质剪接活性。一个具有C-末端谷氨酰胺（Gln），分离自Chilo虹彩病毒（CIV），另一个独特的内含肽，首次在这里报道，具有C-末端天冬氨酸，分离自Carboxydothermus hydrogenoformans（Chy）。在野生型内含肽和N-和C-末端氨基酸取代的几个突变体中检查蛋白质剪接活性。我们证明，这两种野生型内含肽可以蛋白质剪接，可能是由典型的蛋白质剪接机制的新变化。将非典型的C-末端残基替换为典型的Asn，仅在CIV内含肽中保留蛋白质剪接。在Chy内含肽中的所有不同的C-末端取代（Asp（345）至Asn、Gln、Glu和Ala）消除了蛋白质剪接并产生N-和C-末端切割。在以Ala结束的Chy内含肽中观察到的C-末端切割不能通过该残基的环化来解释。我们提出并讨论了几种新的模型，在蛋白质剪接途径的反应。
Inteins are protein-splicing domains present in many proteins. They self-catalyze their excision from the host protein, ligating their former flanks by a peptide bond. The C-terminal residue of inteins is typically an asparagine (Asn). Cyclization of this residue to succinimide causes the final detachment of inteins from their hosts. We studied protein-splicing activity of two inteins with atypical C-terminal residues. One having a C-terminal glutamine (Gln), isolated from Chilo iridescent virus (CIV), and another unique intein, first reported here, with a C-terminal aspartate, isolated from Carboxydothermus hydrogenoformans (Chy). Protein-splicing activity was examined in the wild-type inteins and in several mutants with N- and C-terminal amino acid substitutions. We demonstrate that both wild-type inteins can protein splice, probably by new variations of the typical protein-splicing mechanism. Substituting the atypical C-terminal residue to the typical Asn retained protein-splicing only in the CIV intein. All diverse C-terminal substitutions in the Chy intein (Asp(345) to Asn, Gln, Glu, and Ala) abolished protein-splicing and generated N- and C-terminal cleavage. The observed C-terminal cleavage in the Chy intein ending with Ala cannot be explained by cyclization of this residue. We present and discuss several new models for reactions in the protein-splicing pathway.