Pepinsky R B, Papayannopoulos I A, Chow E P, Krishna N K, Craven R C, Vogt V M
Department of Protein Chemistry, Biogen, Inc., Cambridge, Massachusetts 02142, USA.
J Virol. 1995 Oct;69(10):6430-8. doi: 10.1128/JVI.69.10.6430-6438.1995.
The CA (capsid) protein of avian sarcoma and leukemia viruses occurs in multiple species. Only one form has been previously characterized biochemically. We have now determined that the mature CA protein of avian sarcoma and leukemia viruses exists as three species with different C termini, ending in amino acid residues A-476, A-478, and M-479 of the Gag precursor, respectively. These structures were deduced from a combination of cyanogen bromide peptide mapping, sequence analysis of tryptic peptides, and electrospray mass spectrometry. The three forms of CA were detected in the same ratios in Rous sarcoma virus and avian myeloblastosis virus and therefore are likely to represent a common feature of members of this genus of avian retroviruses. The only previously reported CA species, CAM-479, accounts for only about 36% of the total CA protein, while CAA-476 and CAA-478 account for 55 and 9%, respectively. From the analysis of peptides cleaved in vitro by PR, the viral protease, we infer that the cleavage site between A-476 and A-477 not only is recognized by PR but is the preferred site. We were unable to determine if A-478/A-479 is a cleavage site for PR or alternatively if CAA-478 results from further processing of CAM-479 by a carboxypeptidase. To study the biological significance of residues A-477 to M-479, we constructed genetically altered viruses in which deletions removed either residues 477 to 479 or 477 to 488. The resulting virus particles appeared to assembly with normal efficiencies, but the latter mutant showed slowed proteolytic processing. Neither of the mutants was infectious.
禽肉瘤和白血病病毒的衣壳（CA）蛋白存在多种形式。此前仅有一种形式经过了生化特征鉴定。我们现已确定，禽肉瘤和白血病病毒的成熟CA蛋白以三种具有不同C末端的形式存在，分别在Gag前体的A - 476、A - 478和M - 479氨基酸残基处终止。这些结构是通过溴化氰肽图谱分析、胰蛋白酶肽段序列分析和电喷雾质谱法相结合推导出来的。在劳氏肉瘤病毒和禽成髓细胞瘤病毒中，三种CA形式的检测比例相同，因此它们可能代表了该属禽逆转录病毒成员的一个共同特征。此前唯一报道的CA形式CAM - 479仅占总CA蛋白的约36%，而CAA - 476和CAA - 478分别占55%和9%。通过对病毒蛋白酶PR体外切割的肽段分析，我们推断A - 476和A - 477之间的切割位点不仅能被PR识别，而且是优先切割位点。我们无法确定A - 478/A - 479是否是PR的切割位点，或者CAA - 478是否是CAM - 479被羧肽酶进一步加工的结果。为了研究A - 477至M - 479残基的生物学意义，我们构建了基因改造病毒，其中缺失了477至479或477至488残基。产生的病毒颗粒似乎能以正常效率组装，但后一种突变体的蛋白水解加工过程减缓。这两种突变体均无感染性。
