Nair Smita, Gayathri P, Murthy M R N, Savithri H S
Department of Biochemistry, Indian Institute of Science, Bangalore-560012, Karnataka State, India.
Virology. 2008 Dec 5;382(1):83-90. doi: 10.1016/j.virol.2008.08.034. Epub 2008 Oct 7.
N-terminal serine protease domain of Sesbania mosaic virus polyprotein, requires fused VPg for its activity. W43 of VPg mediates aromatic stacking interactions (characterized by 230 nm positive CD peak) with protease. A stretch of aromatic residues (F269, W271, Y315, and Y319) exposed in the protease domain were mutated to identify the interacting partner of W43. W271A Protease-VPg mutant showed absence of cleavage activity both in vivo and in trans, with concomitant loss of the 230 nm CD peak. F269A Protease-VPg mutant was partially active. Mutations of the tyrosines did not result in loss of protease activity or the CD peak. Interestingly, H275, though not a part of the exposed aromatic stretch, was shown to be essential for protease activity and contributed significantly to the CD peak. Hence, we conclude that W271 and H275 of the protease domain mediate aromatic stacking interactions with W43 of VPg thereby rendering the protease active.
田菁花叶病毒多聚蛋白的N端丝氨酸蛋白酶结构域，其活性需要与融合的VPg结合。VPg的W43介导与蛋白酶的芳香族堆积相互作用（以230nm的正圆二色峰为特征）。对蛋白酶结构域中暴露的一段芳香族残基（F269、W271、Y315和Y319）进行突变，以确定W43的相互作用伙伴。W271A蛋白酶-VPg突变体在体内和体外均无切割活性，同时230nm圆二色峰消失。F269A蛋白酶-VPg突变体具有部分活性。酪氨酸的突变并未导致蛋白酶活性丧失或圆二色峰消失。有趣的是，H275虽然不是暴露的芳香族片段的一部分，但对蛋白酶活性至关重要，并对圆二色峰有显著贡献。因此，我们得出结论，蛋白酶结构域的W271和H275介导与VPg的W43的芳香族堆积相互作用，从而使蛋白酶具有活性。
