Haas A L, Katzung D J, Reback P M, Guarino L A
Department of Biochemistry, Medical College of Wisconsin, Milwaukee 53226, USA.
Biochemistry. 1996 Apr 30;35(17):5385-94. doi: 10.1021/bi9524981.
The marked evolutionary conservation of ubiquitin is assumed to arise from constraints imposed by folding, stability, and interaction of the polypeptide with various components of the ATP, ubiquitin-dependent degradative pathway. The present studies characterize the most divergent (75% identity) of the species-specific ubiquitin isoforms encoded as a late gene product of the baculovirus Autographa californica [Guarino, L. A. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 409-413]. Viral ubiquitin supports 40% of the rate of ATP-dependent degradation exhibited by eukaryotic ubiquitin. Inhibition of proteolysis correlated with a lower steady-state concentration of ubiquitin-conjugated degradative intermediates. Rate studies revealed that viral ubiquitin exerts its effect at the step of isopeptide ligase-catalyzed (E3) ubiquitin conjugation since viral and eukaryotic polypeptides are identical in their abilities to support ATP-coupled activation by E1 and transthiolation to E2 carrier proteins. Other studies demonstrated viral ubiquitin severely attenuated the rate of K48-linked multiubiquitin chain formation in E3-independent conjugation catalyzed by recombination yeast CDC34 or rabbit reticulocyte E232K but not chain elongation of alternate linkages formed by yeast RAD6 or human E2EPF. The latter observations suggest nonconserved positions on viral ubiquitin constitute recognition signals for K48-linked chain formation. Sequence comparison of species-specific ubiquitin isoforms indicates that nonconserved positions localized to a defined region on the polypeptide surface distinct from the basic face required for E1 binding. These results suggest this novel ubiquitin isoform may function in baculoviral replication to block destruction of a short-lived protein(s) by the host degradative pathway, targeted through either E2-catalyzed K48-linked multibiquitin chain formation or general E3-mediated conjugation.
泛素在进化上的显著保守性被认为源于多肽折叠、稳定性以及与ATP泛素依赖性降解途径的各种组分相互作用所施加的限制。本研究对作为杆状病毒苜蓿银纹夜蛾核型多角体病毒（Autographa californica）晚期基因产物编码的物种特异性泛素异构体中差异最大的一种（同一性为75%）进行了表征[瓜里诺，L.A.（1990年）《美国国家科学院院刊》87卷，409 - 413页]。病毒泛素支持真核泛素所表现出的ATP依赖性降解速率的40%。蛋白水解的抑制与泛素缀合降解中间体的较低稳态浓度相关。速率研究表明，病毒泛素在异肽连接酶催化（E3）泛素缀合步骤发挥作用，因为病毒和真核多肽在支持E1介导的ATP偶联激活以及向E2载体蛋白转硫醇化的能力方面是相同的。其他研究表明，病毒泛素严重降低了重组酵母CDC34或兔网织红细胞E232K催化的不依赖E3的缀合中K48连接的多聚泛素链形成速率，但不影响酵母RAD6或人E2EPF形成的交替连接的链延长。后一观察结果表明，病毒泛素上不保守的位置构成了K48连接链形成的识别信号。物种特异性泛素异构体的序列比较表明，不保守的位置定位于多肽表面的一个特定区域，该区域不同于E1结合所需的碱性面。这些结果表明，这种新型泛素异构体可能在杆状病毒复制中发挥作用，通过E2催化的K48连接的多聚泛素链形成或一般的E3介导的缀合来阻止宿主降解途径对一种或多种短命蛋白的破坏。
