Jaskólski M, Miller M, Rao J K, Leis J, Wlodawer A
Crystallography Laboratory, NCI-Frederick Cancer Research and Development Center, Maryland 21701.
Biochemistry. 1990 Jun 26;29(25):5889-98. doi: 10.1021/bi00477a002.
The structure of Rous sarcoma virus protease has been solved by multiple isomorphous replacement in the crystal form belonging to space group P3(1)21, with unit-cell parameters a = 88.95 A and c = 78.90 A. The enzyme belongs to the family of aspartic proteases with two identical subunits composing the active homodimer. The noncrystallographic dyad relating these two subunits was identified after preliminary tracing in the MIR map and was used for phase improvement by electron-density averaging. Structure refinement resulted in a model that included 1772 protein atoms and 252 water molecules, with an R factor of 0.144 for data extending to 2 A. The secondary structure of a retroviral protease molecule closely resembles that of a single domain in pepsin-like aspartic proteases and consists of several beta-strands and of one well-defined and one distorted alpha-helix. The dimer interface is composed of the N- and C-terminal chains from both subunits which are intertwined to form a well-ordered four-stranded antiparallel beta-sheet. In each monomer, the catalytic triad (Asp-Ser-Gly) is located in a loop that forms a part of the psi-structure characteristic to all aspartic proteases. The position of a water molecule between the active-site aspartate residues and the general scheme of H bonding within the active site bear close resemblance to those in pepsin-like aspartic proteases and therefore suggest a similar enzymatic mechanism. The binding cleft over the active site is covered by two flap arms, one from each monomer, which are partially disordered. The retroviral protease dimer has been compared with several enzymes of cellular origin, with chains aligning to an rms deviation of 1.90 A or better.
劳氏肉瘤病毒蛋白酶的结构已通过多重同晶置换法在属于P3(1)21空间群的晶体形式中解析出来，其晶胞参数为a = 88.95 Å和c = 78.90 Å。该酶属于天冬氨酸蛋白酶家族，由两个相同的亚基组成活性同型二聚体。在MIR图中初步追踪后确定了与这两个亚基相关的非晶体学二元轴，并用于通过电子密度平均进行相位改善。结构精修得到了一个包含1772个蛋白质原子和252个水分子的模型，对于延伸至2 Å的数据，R因子为0.144。逆转录病毒蛋白酶分子的二级结构与胃蛋白酶样天冬氨酸蛋白酶中单个结构域的二级结构非常相似，由几条β链和一个明确的α螺旋以及一个扭曲的α螺旋组成。二聚体界面由来自两个亚基的N端和C端链组成，它们相互缠绕形成一个有序的四链反平行β折叠。在每个单体中，催化三联体（天冬氨酸 - 丝氨酸 - 甘氨酸）位于一个环中，该环形成了所有天冬氨酸蛋白酶特有的ψ结构的一部分。活性位点天冬氨酸残基之间水分子的位置以及活性位点内氢键的总体模式与胃蛋白酶样天冬氨酸蛋白酶中的相似，因此表明酶促机制相似。活性位点上方的结合裂隙被两个瓣臂覆盖，每个单体一个，它们部分无序。已将逆转录病毒蛋白酶二聚体与几种细胞来源的酶进行了比较，链的比对均方根偏差为1.90 Å或更小。
