Schwemmle M, Richter M F, Herrmann C, Nassar N, Staeheli P
Abteilung Virologie, Universität Freiburg, Germany.
J Biol Chem. 1995 Jun 2;270(22):13518-23.
MxA is an interferon-induced 76-kDa GTPase that inhibits the multiplication of several RNA viruses. Deleting seven amino acids from the COOH terminus reduced the GTPase activity of purified MxA to 1.4%. MxA mutants with COOH-terminal deletions of 63 or more amino acids lost all ability to hydrolyze GTP and failed to bind guanine nucleotides. By contrast, an MxA deletion mutant consisting of 301 amino acids from the NH2 terminus and 87 amino acids from the COOH terminus retained about 9% of wild-type GTPase activity, underscoring the pivotal role of COOH-terminal sequences. Limited proteolysis of wild-type MxA with proteinase K resulted in two resistant polypeptides of 60 and 10 kDa, respectively, which copurified as a stable complex. The p60-p10 complex exhibited high GTPase activity, suggesting that it included all MxA domains required for this biochemical activity. Sequencing revealed that the NH2 terminus of the 60-kDa polypeptide mapped to leucine 41 and the NH2 terminus of the 10-kDa polypeptide to glutamine 564 of the MxA sequence. Based on these results we propose a model that suggests that the GTP-binding consensus element located in the NH2-terminal half of MxA is held in an active conformation by strong physical interactions with amino acids from the COOH-terminal region.
MxA是一种干扰素诱导的76 kDa GTP酶，可抑制多种RNA病毒的增殖。从COOH末端删除七个氨基酸后，纯化的MxA的GTP酶活性降至1.4%。COOH末端缺失63个或更多氨基酸的MxA突变体丧失了所有水解GTP的能力，并且无法结合鸟嘌呤核苷酸。相比之下，一个由NH2末端的301个氨基酸和COOH末端的87个氨基酸组成的MxA缺失突变体保留了约9%的野生型GTP酶活性，这突出了COOH末端序列的关键作用。用蛋白酶K对野生型MxA进行有限的蛋白水解，分别产生了两个抗性多肽，分子量分别为60 kDa和10 kDa，它们作为稳定的复合物共同纯化。p60-p10复合物表现出高GTP酶活性，表明它包含了这种生化活性所需的所有MxA结构域。测序显示，60 kDa多肽的NH2末端对应于MxA序列中的第41位亮氨酸，10 kDa多肽的NH2末端对应于第564位谷氨酰胺。基于这些结果，我们提出了一个模型，该模型表明位于MxA NH2末端一半的GTP结合共有元件通过与COOH末端区域的氨基酸的强烈物理相互作用而保持在活性构象中。
