A novel chymotrypsin-like component of the multicatalytic proteinase complex optimally active at acidic pH.
A novel chymotrypsin-like component of the multicatalytic proteinase complex optimally active at acidic pH.
复制标题
多催化蛋白酶复合物中的一种新型胰凝乳蛋白酶样成分，在酸性 pH 条件下具有最佳活性。
DOI：
10.1006/abbi.1995.1137
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发表时间：
1995
影响因子：
3.9
通讯作者：
Wilk,S
中科院分区：
文献类型：
--
作者：
Figueiredo-Pereira,ME;Chen,WE;Yuan,HM;Wilk,S
多催化蛋白酶复合物（MPC）或蛋白酶体是一种多聚体，高分子量（700，000），在真核生物和古细菌中发现的溶酶体外蛋白水解酶。它的多个催化位点赋予它对短肽和蛋白质底物的广泛切割特异性。MPC催化活性的最适pH值为中性或弱碱性。我们在这里提出的证据MPC的隐蔽催化成分最佳活性在酸性pH值。研究与疏水性荧光探针提供直接的证据，构象变化所带来的复杂暴露于酸性环境。其中一种新描述的组分，命名为“酸性胰凝乳蛋白酶样活性”，切割底物Boc-Val-Glu-Ala-Leu-2-naphthylamide中的Leu-2-naphthylamide键。与经典的“中性”胰凝乳蛋白酶样活性相比，新描述的组分不受单价阳离子的抑制，并且对中性胰凝乳蛋白酶样活性的抑制剂肽醛Z-Gly-Gly-leucinal不太敏感。此外，我们描述了一种新的有效的肽醛，Z-Ile-GIu（OtBu）-Ala-leucinal的性质，它是MPC的酸性和中性胰凝乳蛋白酶样活性的抑制剂，IC 50值分别为0.25和6.5 μM。在65 μM新合成的肽醛存在下，其他MPC组分如胰蛋白酶样和肽酰-谷氨酰肽水解活性分别仅降低14%和9%。Z-Ile-GIu（OtBu）-Ala-亮氨酸对复合物的胰凝乳蛋白酶样活性的疏水性、效力和特异性使其成为研究MPC的生理作用的有价值的药理学工具。
The multicatalytic proteinase complex (MPC) or proteasome is a multimeric, high-molecular-weight (700,000), extralysosomal proteolytic enzyme found in eukaryotes and in archaebacteria. Its multiple catalytic sites grant it a broad cleavage specificity toward short peptides and protein substrates. The pH optima of the catalytic activities of MPC are in the neutral or slightly alkaline range. We present here evidence for cryptic catalytic components of MPC optimally active at an acidic pH. Studies with a hydrophobic fluorescent probe provide direct evidence for conformational changes brought about by exposing the complex to an acidic environment. One of the newly described components, designated "acidic chymotrypsin-like activity," cleaves the Leu-2-naphthylamide bond in the substrate Boc-Val-Glu-Ala-Leu-2-naphthylamide. Compared with the classical "neutral" chymotrypsin-like activity defined by cleavage of the Leu-p-nitroanilide bond in Z-Gly-Gly-Leu-p-nitroanilide, the newly described component is not inhibited by monovalent cations and is less sensitive to the peptidyl aldehyde Z-Gly-Gly-leucinal, an inhibitor of the neutral chymotrypsin-like activity. In addition, we describe the properties of a novel potent peptidyl aldehyde, Z-Ile-GIu(OtBu)-Ala-leucinal, which is an inhibitor of both the acidic and neutral chymotrypsin-like activities of MPC, with IC50values of 0.25 and 6.5 μM, respectively. In the presence of 65 μM of the newly synthesized peptidyl aldehyde, other MPC components such as the trypsin-like and peptidyl-glutamyl peptide hydrolyzing activities were decreased only by 14 and 9%, respectively. The hydrophobicity, potency, and specificity of Z-Ile-GIu(OtBu)-Ala-leucinal toward the chymotrypsin-like activities of the complex make it a valuable pharmacological tool with which to investigate the physiological roles of MPC.