Proteolytic action of GlpG, a rhomboid protease in the Escherichia coli cytoplasmic membrane
Proteolytic action of GlpG, a rhomboid protease in the Escherichia coli cytoplasmic membrane
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DOI：
10.1021/bi051363k
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发表时间：
2005-10-18
期刊：
影响因子：
2.9
通讯作者：
Akiyama, Y
中科院分区：
文献类型：
--
作者：
Maegawa, S;Ito, K;Akiyama, Y
我们的特点是大肠杆菌GlpG作为一种膜包埋蛋白酶和一个可能的球员在这种生物体的调节膜内蛋白水解。从序列特征来看，它属于广泛保守的菱形膜蛋白酶家族。我们验证了预期的GlpG拓扑结构，它穿过膜六次。发现具有N-末端和周质定位的β-内酰胺酶（Bla）结构域、LacY衍生的跨膜区和胞质麦芽糖结合蛋白（MBP）成熟结构域的模型蛋白在体内被GlpG依赖性切割。使用纯化的GlpG和纯化的模型底物蛋白质在体外再现这种蛋白水解反应，并且所示的切割发生在Ser和Asp之间的高局部亲水性区域中，该区域可能位于质膜中而不是膜内位置。GlpG的保守Ser和His残基是其蛋白水解活性所必需的。我们的研究结果使用几种不同形式的模型蛋白表明，GlpG识别功能的跨膜区的基板。这些结果指出了这类有趣的膜嵌入蛋白酶的详细分子机制和细胞分析。
We characterized Escherichia coli GlpG as a membrane-embedded protease and a possible player in the regulated intramembrane proteolysis in this organism. From the sequence features, it belong's to the widely conserved rhomboid family of membrane proteases. We verified the expected topology of GlpG, and it traverses the membrane six times. A model protein having an N-terminal and periplasmically localized beta-lactamase (Bla) domain, a LacY-derived transmembrane region, and a cytosolic maltose binding protein (MBP) mature domain was found to be GlpG-dependently cleaved in vivo. This proteolytic reaction was reproduced in vitro using purified GlpG and purified model substrate protein, and the cleavage was shown to occur between Ser and Asp in a region of high local hydrophilicity, which might be located in a juxtamembrane rather than an intramembrane position. The conserved Ser and His residues of GlpG were essential for the proteolytic activities. Our results using several variant forms of the model protein suggest that GlpG recognizes features of the transmembrane regions of substrates. These results point to a detailed molecular mechanism and cellular analysis of this interesting class of membrane-embedded proteases.