Maxwell S A, Arlinghaus R B
J Gen Virol. 1985 Jan;66 ( Pt 1):97-107. doi: 10.1099/0022-1317-66-1-97.
In vitro proteolytic cleavage of the Gazdar murine sarcoma virus (Gz-MuSV) p65gag polypeptide (Gz-p65gag) was facilitated by detergent-disrupted Moloney murine leukaemia virus (Mo-MuLV). Incubation of radioactively labelled Gz-p65gag in the presence of unlabelled Mo-MuLV under optimal conditions resulted in the cleavage of Gz-p65gag to proteins of 40000 (P40) and 25000 (P25) Mr. P40 and P25 appeared to be similar in both mobility and antigenicity to Mo-MuLV intermediates, Pr40gag and Pr25gag, previously found in infected cells. Additional proteins of 30000 (Gz-p30), 15000 (Gz-p12), 12000 (Gz-p15) and 10000 (Gzp10) Mr were also generated upon cleavage of Gz-p65gag and contained antigenic determinants of Mo-MuLV structural proteins p30, pp12, p15 and p10, respectively. Both detergent-disrupted Mo-MuLV and Rauscher murine leukaemia virus produced similar cleavage profiles. Trypsin and detergent-disrupted mouse mammary tumour virus generated cleavage patterns very different from that produced by Mo-MuLV. Both visual and quantitative time studies of the reaction indicated that P40 gave rise to Gz-p30 and Gz-p10. Tryptic peptide mapping of Gz-p65gag and its cleavage products supported the results obtained from both immunoprecipitation studies with anti-gag sera and the kinetics of cleavage of Gz-p65gag. Both Mo-MuLV Pr65gag and Gz-p65gag were found to be very similar in primary sequence as judged by peptide mapping. P40 produced tryptic peptides that co-migrated with Mo-MuLV p30 peptides; P25 contained tryptic peptides that were also found in Mo-MuLV p15. Gz-p30 and Gz-p15 contained the tryptic peptides of Mo-MuLV p30 and p15, respectively, that were found in P40 and P25. The Gz-p10 fraction contained a tryptic peptide that was also found in P40, but not p30. These results provide good evidence that the protease packaged within Mo-MuLV can cleave, in vitro, the gag-related polyprotein of Gz-MuSV in a manner very similar to the processing of Mo-MuLV Pr65gag in infected cell culture systems.
去污剂破坏的莫洛尼氏鼠白血病病毒（Mo-MuLV）促进了体外对加兹达尔鼠肉瘤病毒（Gz-MuSV）p65gag多肽（Gz-p65gag）的蛋白水解切割。在最佳条件下，将放射性标记的Gz-p65gag与未标记的Mo-MuLV一起孵育，导致Gz-p65gag被切割成分子量为40000（P40）和25000（P25）的蛋白质。P40和P25在迁移率和抗原性方面似乎与先前在感染细胞中发现的Mo-MuLV中间体Pr40gag和Pr25gag相似。切割Gz-p65gag后还产生了分子量为30000（Gz-p30）、15000（Gz-p12）、12000（Gz-p15）和10000（Gzp10）的其他蛋白质，它们分别含有Mo-MuLV结构蛋白p30、pp12、p15和p10的抗原决定簇。去污剂破坏的Mo-MuLV和劳舍尔氏鼠白血病病毒产生了相似的切割图谱。胰蛋白酶和去污剂破坏的小鼠乳腺肿瘤病毒产生的切割模式与Mo-MuLV产生的非常不同。对该反应的视觉和定量时间研究均表明，P40产生了Gz-p30和Gz-p10。Gz-p65gag及其切割产物的胰蛋白酶肽图谱支持了用抗gag血清进行免疫沉淀研究以及Gz-p65gag切割动力学所获得的结果。通过肽图谱判断，发现Mo-MuLV Pr65gag和Gz-p65gag在一级序列上非常相似。P40产生的胰蛋白酶肽与Mo-MuLV p30肽共迁移；P25含有的胰蛋白酶肽也存在于Mo-MuLV p15中。Gz-p30和Gz-p15分别含有在P40和P25中发现的Mo-MuLV p30和p15的胰蛋白酶肽。Gz-p10部分含有一种也存在于P40中但不存在于p30中的胰蛋白酶肽。这些结果提供了充分的证据，表明包装在Mo-MuLV内的蛋白酶在体外能够以与感染细胞培养系统中Mo-MuLV Pr65gag的加工方式非常相似的方式切割Gz-MuSV的gag相关多蛋白。
