Huang L, Shen M, Chernushevich I, Burlingame A L, Wang C C, Robertson C D
Department of Pharmaceutical Chemistry, University of California, San Francisco 94143-0446, USA.
Mol Biochem Parasitol. 1999 Aug 20;102(2):211-23. doi: 10.1016/s0166-6851(99)00096-1.
We have determined peptide sequences of three Trypanosoma brucei proteasome subunit proteins by mass spectrometry of tryptic digests of the proteins purified by two-dimensional (2-D) polyacrylamide gel electrophoresis. Three genes identified by the sequence of their cDNA encode the peptides identified in these three proteins. The three proteins predicted from the gene sequences have significant similarity to other known proteasome subunits and represent an alpha6 type subunit (TbPSA6), and two beta-type subunits belonging to the beta1-type (TbPSB1) and beta2 type (TbPSB2). The sequences of both beta-subunits predict formation of catalytically active subunits through proteolytic processing. The prediction is supported by the presence in each of the two beta-subunits of a tryptic peptide that has the correctly processed N-terminus that creates the threonine nucleophile of the mature protein. This peptide cannot be generated by trypsin because of the required cleavage of a glycine-threonine bond. It is thus likely that there are at least two catalytically active beta-subunits, TbPSB1 and TbPSB2, present in the mature 20S proteasome from T. brucei.
我们通过对二维（2-D）聚丙烯酰胺凝胶电泳纯化的蛋白质进行胰蛋白酶消化后的质谱分析，确定了三种布氏锥虫蛋白酶体亚基蛋白的肽序列。根据其cDNA序列鉴定出的三个基因编码了在这三种蛋白质中鉴定出的肽。从基因序列预测的这三种蛋白质与其他已知的蛋白酶体亚基具有显著相似性，分别代表一个α6型亚基（TbPSA6）以及两个属于β1型（TbPSB1）和β2型（TbPSB2）的β型亚基。两个β亚基的序列预测通过蛋白水解加工形成催化活性亚基。这一预测得到了支持，因为在两个β亚基中的每一个中都存在一个胰蛋白酶肽，其具有经过正确加工的N端，该N端形成了成熟蛋白的苏氨酸亲核试剂。由于需要切割甘氨酸-苏氨酸键，胰蛋白酶无法产生这种肽。因此，在来自布氏锥虫的成熟20S蛋白酶体中可能至少存在两种具有催化活性的β亚基，即TbPSB1和TbPSB2。
