Dual specificities of the glyoxysomal/peroxisomal processing protease Deg15 in higher plants
Dual specificities of the glyoxysomal/peroxisomal processing protease Deg15 in higher plants
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DOI：
10.1073/pnas.0704733104
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发表时间：
2007-07-03
影响因子：
11.1
通讯作者：
Gietl, Christine
中科院分区：
文献类型：
--
作者：
Helm, Michael;Lueck, Carsten;Gietl, Christine
乙醛酸酶体是参与脂质动员的过氧化物酶体的一个亚类。定义了两种不同的过氧化物酶体靶向信号（PTS），C-末端PTS 1和N-末端PTS 2。PTS 2对蛋白质输入的加工在高等真核生物中是保守的。切割位点通常在P1或P2处含有Cys。采用柱层析、制备性等电聚焦和2DPAGE等方法从西瓜（Citrullus vulgaris）贮藏脂肪的子叶中分离纯化了乙醛酸酶体加工蛋白酶（GPP）。GPP以两种形式出现，72-kDa单体和144-kDa二聚体，它们彼此平衡。的平衡是移动对Ca 2+去除朝向单体和对Ca 2+除了朝向二聚体。该单体是一种通用的降解蛋白酶，并被变性蛋白质激活。由于证明单体的底物特异性（Phi-Arg/Lys向下箭头）不同于单体和二聚体混合物的加工底物特异性（Cys-Xxx向下箭头/Xxx-Cys向下箭头），因此Climer构成加工蛋白酶。拟南芥基因组分析揭示了三种蛋白酶，预测是在过氧化物酶体，Deg-protease，pitrilysin-like金属肽酶，和Lonprotease。针对来自拟南芥的过氧化物酶体Degprotease（Deg 15）的特异性抗体将西瓜GPPs鉴定为Deg 15。拟南芥DEG 15基因中的敲除突变（At 1g 28320）阻止将乙醛酸脱氢酶前体加工成成熟形式。因此，GPP/Deg 15属于以大肠杆菌DegP为原型的胰蛋白酶样丝氨酸蛋白酶组。然而，GPP/Deg 15具有特定的特性，因此是Deg 15蛋白酶中的一个新亚组。
Glyoxysomes are a subclass of peroxisomes involved in lipid mobilization. Two distinct peroxisomal targeting signals (PTSs), the C-terminal PTS1 and the N-terminal PTS2, are defined. Processing of the PTS2 on protein import is conserved in higher eukaryotes. The cleavage site typically contains a Cys at P1 or P2. We purified the glyoxysomal processing protease (GPP) from the fat-storing cotyledons of watermelon (Citrullus vulgaris) by column chromatography, preparative native isoelectric focusing, and 2D PAGE. The GPP appears in two forms, a 72-kDa monomer and a 144-kDa dimer, which are in equilibrium with one another. The equilibrium is shifted on Ca2+ removal toward the monomer and on Ca2+ addition toward the dimer. The monomer is a general degrading protease and is activated by denatured proteins. The climer constitutes the processing protease because the substrate specificity proven for the monomer (Phi-Arg/Lys down arrow) is different from the processing substrate specificity (Cys-Xxx down arrow /Xxx-Cys down arrow) found with the mixture of monomer and dimer. The Arabidopsis genome analysis disclosed three proteases predicted to be in peroxisomes, a Deg-protease, a pitrilysin-like metallopeptidase, and a Lonprotease. Specific antibodies against the peroxisomal Degprotease from Arabidopsis (Deg15) identify the watermelon GPPas a Deg15. A knockout mutation in the DEG15 gene of Arabidopsis (At1g28320) prevents processing of the glyoxysomal malate dehydrogenase precursor to the mature form. Thus, the GPP/Deg15 belongs to a group of trypsin-like serine proteases with Escherichia coli DegP as a prototype. Nevertheless, the GPP/Deg15 possesses specific characteristics and is therefore a new subgroup within the Deg proteases.