原核生物蛋白質體中富含甘胺酸與精胺酸基序之分析

Glycine and arginine rich (GAR) motif為蛋白質中含有RG或RGG重複序列的區域。為了系統化分析蛋白質中的GAR motif，我們實驗室發展了GMF軟體，可以於輸入的序列中界定並列出GAR特徵。在真核生物中，GAR motif的精胺酸常發生甲基化，這種轉譯後修飾在轉錄調控、RNA剪接、DNA修復與細胞訊號傳遞中扮演關鍵角色。相較之下，原核生物缺乏這類轉譯後修飾。本研究旨在探討古菌與細菌中GAR motif是否在特徵上與真核生物有所不同。我們從NCBI取得在先前研究中用於建構演化樹的各物種完整蛋白質體，並以GMF進行分析。與真核生物相比，古菌與細菌表現出以下特徵：含GAR motif的蛋白比例較低、GAR motif中非RG的胺基酸占比大於50%的蛋白比例較高、RGG / RG比值較低、GAR motif較短，以及單一蛋白中所含的GAR motif數量較少。在古菌與細菌中，最常見含GAR motif的蛋白包括ABC transporter ATP-binding protein、GNAT family N-acetyltransferase、MFS transporter、DEAD / DEAH box helicase、ATP-binding protein。此外，我們也分析了RG或RGG重複次數較高的蛋白質。針對特定蛋白質，我們進行多重序列比對以觀察GAR motif的保守性，或利用BLAST分析在其他物種中的同源蛋白是否也具有GAR motif。例如DEAD / DEAH box解旋酶擁有保守的DEAD motif，但其GAR motif通常不具有保守性，且數量與長度變異較大。在古菌中，某些NifB / NifX family molybdenum-iron cluster-binding protein含有超長的GAR motif，而其中編碼GAR motif的區域可能參與G-quadruplex結構的形成。在細菌物種樹中，基因序列中GC含量較高的放線菌門比起GC含量較低的厚壁菌門，擁有更高比例的GAR motif蛋白，且每個蛋白中GAR motif的數量也更多，顯示高GC含量可能有助於形成更多的GAR motif。此外，我們也進行基因本體（GO）分析，以進一步了解含GAR motif蛋白的潛在生物功能。本研究應有助於了解GAR motif在不同功能蛋白質中扮演的角色，以及其在各類生物系統中的演化。

The glycine and arginine rich (GAR) motif refers to regions in proteins containing repeated RG or RGG sequences. To systematically analyze GAR motifs in proteins, our laboratory developed the GMF software, which identifies and lists GAR features in input sequences. In eukaryotes, arginine residues within GAR motifs are frequently methylated—a post-translational modification that plays critical roles in transcriptional regulation, RNA splicing, DNA repair, and cell signaling. In contrast, such modifications are largely absent in prokaryotes. This study aims to investigate whether the characteristics of GAR motifs in archaea and bacteria differ from those in eukaryotes. We obtained complete proteomes of various species—previously used for constructing phylogenetic trees—from NCBI and analyzed them using GMF. Compared to eukaryotes, archaea and bacteria exhibit following features: a lower proportion of proteins containing GAR motifs; a higher proportion of proteins where over 50% of the GAR motif residues are non-RG amino acids; a lower RGG / RG ratio; shorter GAR motifs; and fewer GAR motifs per protein. In archaea and bacteria, the most common GAR-containing proteins include ABC transporter ATP-binding proteins, GNAT family N-acetyltransferases, MFS transporters, DEAD / DEAH box helicases, and ATP-binding proteins. We also analyzed proteins with high counts of RG or RGG repeats. For selected proteins, multiple sequence alignment was performed to assess GAR motif conservation, and BLAST was used to investigate whether homologs in other species also possess GAR motifs. For example, although DEAD / DEAH box helicases have conserved DEAD motifs, their GAR motifs are typically not conserved and vary greatly in number and length. In archaea, some NifB / NifX family molybdenum-iron cluster-binding proteins contain exceptionally long GAR motifs, and the regions encoding these motifs may contribute to G-quadruplex structure formation. Among bacterial phyla, Actinomycetota—known for their high GC content—have a higher proportion of GAR motif-containing proteins and more GAR motifs per protein compared to low-GC Bacillota, suggesting that high GC content may promote GAR motif formation. Finally, we conducted Gene Ontology (GO) analysis to further explore the potential biological functions of GAR motif-containing proteins. This study contributes to a better understanding of the roles GAR motifs play in various functional proteins and their evolutionary dynamics across biological systems.