Ni R, Tomita Y, Tokunaga F, Liang T J, Noda C, Ichihara A, Tanaka K
Institute for Enzyme Research, University of Tokushima, Japan.
Biochim Biophys Acta. 1995 Oct 17;1264(1):45-52. doi: 10.1016/0167-4781(95)00113-u.
A new subunit, named RC6-I, of the rat 20 S proteasome was purified and the partial amino acid sequences of several peptide fragments obtained by digestion with lysyl-endopeptidase were determined by Edman degradation. Amplification of cDNAs encoding RC6-I by the polymerase chain reaction (PCR) technique revealed two types of cDNA, tentatively designated as RC6-IL and RC6-IS in order of size. The nucleotide sequences of the two cDNAs are identical except that RC6-IL contains an insertion of 18 nucleotides in the coding region compared with RC6-IS. The polypeptide predicted from the open reading frame of RC6-IS cDNA consists of 248 amino acid residues with a calculated molecular weight of 27,783. These values are consistent with those obtained by protein chemical analyses. Computer-assisted homology analysis showed that RC6-I belongs to the alpha-type subfamily of the proteasome gene family, which shows similarity to the alpha-subunit of the archaebacterium Thermoplasma acidophilum proteasome, and that the 18 nucleotide insert, encoding six amino acid residues, VVASVS, appears to be unique to RC6-IL, because this motif has not been conserved in any other alpha-type subunit. By reverse transcription (RT)-PCR analysis, the mRNAs for both RC6-IL and RC6-IS were found in all the rat tissues examined. These results suggest that proteasomes are present as a heterogeneous population, possibly for acquisition of diversity of functions.
纯化了大鼠20S蛋白酶体的一个新亚基，命名为RC6-I，并通过埃德曼降解法测定了用赖氨酰内肽酶消化得到的几个肽片段的部分氨基酸序列。通过聚合酶链反应（PCR）技术扩增编码RC6-I的cDNA，发现了两种类型的cDNA，根据大小顺序暂定为RC6-IL和RC6-IS。这两种cDNA的核苷酸序列相同，只是与RC6-IS相比，RC6-IL在编码区有一个18个核苷酸的插入。从RC6-IS cDNA的开放阅读框预测的多肽由248个氨基酸残基组成，计算分子量为27783。这些值与蛋白质化学分析得到的值一致。计算机辅助同源性分析表明，RC6-I属于蛋白酶体基因家族的α型亚家族，与嗜热栖热放线菌蛋白酶体的α亚基相似，并且编码六个氨基酸残基VVASVS的18个核苷酸插入似乎是RC6-IL特有的，因为这个基序在任何其他α型亚基中都没有保守。通过逆转录（RT）-PCR分析，在所有检测的大鼠组织中都发现了RC6-IL和RC6-IS的mRNA。这些结果表明蛋白酶体以异质群体的形式存在，可能是为了获得功能的多样性。
