Luo Jiann-Kae, Hornby Judith A T, Wallace Louise A, Chen Jihong, Armstrong Richard N, Dirr Heini W
University Research Council Protein Structure-Function Research Programme, School of Molecular and Cell Biology, University of the Witwatersrand, Johannesburg 2050, South Africa.
Protein Sci. 2002 Sep;11(9):2208-17. doi: 10.1110/ps.0208002.
Rat micro class glutathione transferases M1-1 and M2-2 are homodimers that share a 78% sequence identity but display differences in stability. M1-1 is more stable at the secondary and tertiary structural levels, whereas its quaternary structure is less stable. Each subunit in these proteins consists of two structurally distinct domains with intersubunit contacts occurring between domain 1 of one subunit and domain 2 of the other subunit. The chimeric subunit variants M(12), which has domain 1 of M1 and domain 2 of M2, and its complement M(21), were used to investigate the conformational stability of the chimeric homodimers M(12)-(12) and M(21)-(21) to determine the contribution of each domain toward stability. Exchanging entire domains between class micro GSTs is accommodated by the GST fold. Urea-induced equilibrium unfolding data indicate that whereas the class micro equilibrium unfolding mechanism (i.e., N(2) <--> 2I <--> 2U) is not altered, domain exchanges impact significantly on the conformational stability of the native dimers and monomeric folding intermediates. Data for the wild-type and chimeric proteins indicate that the order of stability for the native dimer (N(2)) is M2-2 > M(12)-(12) M1-1 approximately M(21)-(21), and that the order of stability of the monomeric intermediate (I) is M1 > M2 approximately M(12) > M(21). Interactions involving Arg 77, which is topologically conserved in GSTs, appear to play an important role in the stability of both the native dimeric and folding monomeric structures.
大鼠微粒体谷胱甘肽转移酶M1-1和M2-2是同型二聚体，它们的序列同一性为78%，但在稳定性上存在差异。M1-1在二级和三级结构水平上更稳定，而其四聚体结构则不太稳定。这些蛋白质中的每个亚基都由两个结构不同的结构域组成，亚基间的接触发生在一个亚基的结构域1和另一个亚基的结构域2之间。嵌合亚基变体M(12)（具有M1的结构域1和M2的结构域2）及其互补体M(21)，用于研究嵌合同型二聚体M(12)-(12)和M(21)-(21)的构象稳定性，以确定每个结构域对稳定性的贡献。微粒体类谷胱甘肽转移酶之间整个结构域的交换可被谷胱甘肽转移酶折叠所容纳。尿素诱导的平衡去折叠数据表明，虽然微粒体类平衡去折叠机制（即N(2) <--> 2I <--> 2U）没有改变，但结构域交换对天然二聚体和单体折叠中间体的构象稳定性有显著影响。野生型和嵌合蛋白的数据表明，天然二聚体（N(2)）的稳定性顺序为M2-2 > M(12)-(12) > M1-1 ≈ M(21)-(21)，单体中间体（I）的稳定性顺序为M1 > M2 ≈ M(12) > M(21)。涉及精氨酸77的相互作用（在谷胱甘肽转移酶中拓扑保守）似乎在天然二聚体和折叠单体结构的稳定性中都起着重要作用。
