Mapping N- to C-terminal allosteric coupling through disruption of a putative CD74 activation site in D-dopachrome tautomerase.
Mapping N- to C-terminal allosteric coupling through disruption of a putative CD74 activation site in D-dopachrome tautomerase.
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DOI：
10.1016/j.jbc.2023.104729
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发表时间：
2023-06
影响因子：
4.8
通讯作者：
Lisi, George P
中科院分区：
文献类型：
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作者：
Chen, Emily;Widjaja, Vinnie;Kyro, Gregory;Allen, Brandon;Das, Pragnya;Prahaladan, Varsha M;Bhandari, Vineet;Lolis, Elias J;Batista, Victor S;Lisi, George P
巨噬细胞移动抑制因子（MIF）蛋白家族由MIF和D-多巴色素互变异构酶（也称为MIF-2）组成。这些同源物共有34%的序列同一性，同时保持几乎不可区分的三级和四级结构，这可能是其重叠功能的主要贡献者，包括分化簇74（CD 74）受体的结合和活化以介导炎症。在此之前，我们研究了一种新的变构位点，Tyr 99，通过动态偶联残基的“途径”调节MIF的N-末端催化活性。在一项比较研究中，我们揭示了一个类似的变构途径在MIF-2，尽管其独特的一级序列。MIF和MIF-2 N末端的破坏也减少了C末端的CD 74活化，尽管在MIF-2中受体活化位点并不完全确定。在本研究中，我们使用定点突变，核磁共振波谱，分子模拟，在体外和体内生物化学探索推定的CD 74活化区的MIF-2的基础上同源性的MIF。我们还证实了它的相互结构耦合的MIF-2变构位点和N-末端酶位点。因此，我们提供了进一步的了解CD 74活化位点的MIF-2和它的变构偶联免疫调节。
The macrophage migration inhibitory factor (MIF) protein family consists of MIF and D-dopachrome tautomerase (also known as MIF-2). These homologs share 34% sequence identity while maintaining nearly indistinguishable tertiary and quaternary structure, which is likely a major contributor to their overlapping functions, including the binding and activation of the cluster of differentiation 74 (CD74) receptor to mediate inflammation. Previously, we investigated a novel allosteric site, Tyr99, that modulated N-terminal catalytic activity in MIF through a “pathway” of dynamically coupled residues. In a comparative study, we revealed an analogous allosteric pathway in MIF-2 despite its unique primary sequence. Disruptions of the MIF and MIF-2 N termini also diminished CD74 activation at the C terminus, though the receptor activation site is not fully defined in MIF-2. In this study, we use site-directed mutagenesis, NMR spectroscopy, molecular simulations, in vitro and in vivo biochemistry to explore the putative CD74 activation region of MIF-2 based on homology to MIF. We also confirm its reciprocal structural coupling to the MIF-2 allosteric site and N-terminal enzymatic site. Thus, we provide further insight into the CD74 activation site of MIF-2 and its allosteric coupling for immunoregulation.