A 26-residue peptide possessing the αN-helix motif of the protein kinase A (PKA) regulatory subunit-like proteins from the Trypanozoom subgenera (VAP26, sequence = VAPYFEKSEDETALILKLLTYNVLFS), was shown to inhibit the enzymatic activity of the Trypanosoma equiperdum PKA catalytic subunit-like protein, in a similar manner that the mammalian heat-stable soluble PKA inhibitor known as PKI. However, VAP26 does not contain the PKI inhibitory sequence. Bioinformatics analyzes of the αN-helix motif from various Trypanozoon PKA regulatory subunit-like proteins suggested that the sequence could form favorable peptide-protein interactions of hydrophobic nature with the PKA catalytic subunit-like protein, which possibly may represent an alternative PKA inhibitory mechanism. The sequence of the αN-helix motif of the Trypanozoon proteins was shown to be highly homologous but significantly divergent from the corresponding αN-helix motifs of their Leishmania and mammalian counterparts. This sequence divergence contrasted with the proposed secondary structure of the αN-helix motif, which appeared conserved in every analyzed regulatory subunit-like protein. In silico mutation experiments at positions I234, L238 and F244 of the αN-helix motif from the Trypanozoon proteins destabilized both the specific motif and the protein. On the contrary, mutations at positions T239 and Y240 stabilized the motif and the protein. These results suggested that the αN-helix motif from the Trypanozoon proteins probably possessed a different evolutionary path than their Leishmania and mammalian counterparts. Moreover, finding stabilizing mutations indicated that new inhibitory peptides may be designed based on the αN-helix motif from the Trypanozoon PKA regulatory subunit-like proteins.

Graphical Abstract

中文翻译：

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能够抑制马锥虫蛋白激酶 A 催化亚基样蛋白酶活性的新型肽的分析

来自锥虫亚属（VAP26，序列 = VAPYFEKSEDETALILKLLTYNVLFS ）的蛋白激酶 A (PKA) 调节亚基样蛋白的 αN 螺旋基序的 26 残基肽，被证明可以抑制马匹锥虫PKA 催化亚基的酶活性类蛋白，其作用方式与哺乳动物热稳定可溶性 PKA 抑制剂（称为 PKI）类似。然而，VAP26 不包含 PKI 抑制序列。对各种锥虫PKA 调节亚基样蛋白的 αN 螺旋基序进行的生物信息学分析表明，该序列可以与 PKA 催化亚基样蛋白形成有利的疏水性肽-蛋白相互作用，这可能代表另一种 PKA 抑制机制。锥虫蛋白的 αN-螺旋基序序列被证明与利什曼原虫和哺乳动物对应物的相应 αN-螺旋基序高度同源，但显着不同。这种序列差异与所提出的 αN 螺旋基序的二级结构形成鲜明对比，后者在每个分析的调节亚基样蛋白中似乎都是保守的。在计算机模拟突变实验中，锥虫蛋白αN 螺旋基序的 I234、L238 和 F244 位点使特定基序和蛋白质都不稳定。相反，T239 和 Y240 位点的突变稳定了基序和蛋白质。这些结果表明，锥虫蛋白的 αN 螺旋基序可能具有与利什曼原虫和哺乳动物对应物不同的进化路径。此外，发现稳定突变表明可以基于锥虫PKA 调节亚基样蛋白的 αN 螺旋基序设计新的抑制肽。

图形概要