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首页> 外文期刊>Journal of Molecular Biology >Crystal structure of Bacillus subtilis signal peptide peptidase A
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Crystal structure of Bacillus subtilis signal peptide peptidase A

机译：枯草芽孢杆菌信号肽肽酶A的晶体结构

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Signal peptide peptidase A (SppA) is a membrane-bound self- compartmentalized serine protease that functions to cleave the remnant signal peptides left behind after protein secretion and cleavage by signal peptidases. SppA is found in plants, archaea and bacteria. Here, we report the first crystal structure of a Gram-positive bacterial SppA. The 2.4-?-resolution structure of Bacillus subtilis SppA (SppA BS) catalytic domain reveals eight SppA BS molecules in the asymmetric unit, forming a dome-shaped octameric complex. The octameric state of SppA BS is supported by analytical size-exclusion chromatography and multi-angle light scattering analysis. Our sequence analysis, mutagenesis and activity assays are consistent with Ser147 serving as the nucleophile and Lys199 serving as the general base; however, they are located in different region of the protein, more than 29 ? apart. Only upon assembling the octamer do the serine and lysine come within close proximity, with neighboring protomers each providing one-half of the catalytic dyad, thus producing eight separate active sites within the complex, twice the number seen within Escherichia coli SppA (SppA EC). The SppA BS S1 substrate specificity pocket is deep, narrow and hydrophobic, but with a polar bottom. The S3 pocket, which is constructed from two neighboring proteins, is shallower, wider and more polar than the S1 pocket. A comparison of these pockets to those seen in SppA EC reveals a significant difference in the size and shape of the S1 pocket, which we show is reflected in the repertoire of peptides the enzymes are capable of cleaving.
机译：信号肽肽酶A（SppA）是一种与膜结合的自间隔丝氨酸蛋白酶，其功能是切割蛋白质分泌和信号肽酶切割后留下的残余信号肽。 SppA存在于植物，古细菌和细菌中。在这里，我们报告革兰氏阳性细菌SppA的第一个晶体结构。枯草芽孢杆菌SppA（SppA BS）催化域的2.4-α分辨率结构揭示了不对称单元中的八个SppA BS分子，形成了圆顶形的八聚体。分析尺寸排阻色谱法和多角度光散射分析法支持SppA BS的八聚体状态。我们的序列分析，诱变和活性分析与作为亲核试剂的Ser147和作为一般碱基的Lys199一致;但是，它们位于蛋白质的不同区域，超过29个？分开。只有在组装八聚体后，丝氨酸和赖氨酸才会紧密靠近，相邻的protomer各自提供一半的催化二倍体，从而在复合物中产生八个独立的活性位点，是大肠杆菌SppA（SppA EC）中见到的两倍。 SppA BS S1底物特异性口袋深，窄且疏水，但底部为极性。由两个相邻蛋白质构成的S3口袋比S1口袋更浅，更宽且更具极性。将这些口袋与在SppA EC中看到的口袋进行比较后发现，S1口袋的大小和形状存在显着差异，这显示在酶能够裂解的肽库中。

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