The new mechanism of pathogen ubiture modification is explained

June 2, 2020, Nature Communications published the latest research results from the
Institute of Biophysics's Gao Wei Research Group online. The work analyzed the complex structure of the new ubiganized enzyme MavC and the substrate protein UPE2N and ubibin molecules from the highly pathogenic Legionella pneumophilus, and the two complex structures of MavC and its homoglobin MvcA and negative regulatory factor Lpg2149. Combined with a large number of bio-chemical and cellular experiments, the molecular mechanism of catalysis and regulation of this new ubiganized modification system is expounded.
protein ubibination modification is one of the most important post-translation modifications of the nucleocytes. Classic ubibin modifications need to be catalyzed by E1-E2-E3 trienzyme cascading reactions, while the recently identified two types of L. The pneumophila effect protein (SidE and MavC) can catalyz the ubibin modification of the host protein in a new way that does not rely on E1-E2-E3 (Figure 1).
team reported on the structure and molecular mechanisms of the SidE family protein catalytic reaction in Cell in 2018, and the latest work focuses on the MavC family protein.MavC's genomic cluster encodes three effect proteins: Lpg2147 (MavC), Lpg2148 (MavC paralog A or MvcA) and Lpg2149. Both MavC and MvcA can catalyz the deamination reaction of ubibin molecules (Ub-Q40 to Ub-E40), while Lpg2149 inhibits the activity of both deaminases. Recent studies have shown that MavC can also specifically ubigen modify the host protein UBUE2N, thereby inhibiting the normal E2 function of UBUE2N, thereby inhibiting the activity of the NF-B path. Interestingly, although MvcA is very similar to MavC, MvcA does not catalytic UBA2N ubibination. Using MavC catalysis, the researchers prepared ubiminated modified UBE2N (UBE2N-Ub) and analyzed the crystal structure of the MavC and UBE2N-Ub complexes (Figure 2). The structure directly reveals the interface between MavC and UBE2N and Ub, and shows that MavC uses the same active center to catalytic deamine reaction and ubimination reaction.
The sequence and structure comparison between MvcA and MavC shows that the presence of key amino acids that bind to UVE2N is not conservative in MvcA's Insertion domain, which explains why MvcA is unable to stabilize the binding of UME2N in free state, and why UBUE2N's ubiquitin modification cannot be catalyticized. These structurally based conclusions have also been further validated through bio-chemical and cellular experiments.unexpectedly, MavC and MvcA also have the activity of de-ubibinase, which specifically removes mavC-mediated UME2N ubially modified. Since MvcA is more active than MavC's de-ubibination, MavC and MvcA are likely to perform the functions of ubibinase and de-ubibinase, respectively, in actual infection. The findings suggest that MavC can use the same active pocket to catalyz the three enzymatic reactions of deamining, ubimination, and de-ubibinization.
further analyzed the crystal structure of the MavC-Lpg2149 complex and the MvcA-Lpg2149 complex (Figure 3). Unlike the crystal structure in the free state, Lpg2149 interacts with MavC or MvcA in a monomer form. Since Lpg2149 and Ub occupy similar binding positions, Lpg2149 can directly interfere with the interaction between Ub and MavC/MvcA. Functional experiments show that Lpg2149 can indeed inhibit deamining, ubimination and de-ubimination at the same time. Interestingly, the free-state MavC itself has a binding composition of Lpg2149, while MvcA requires a large configuration change to achieve a combination with Lpg2149. Further combined experiments showed that MavC-Lpg2149 had an affinity about 50 times stronger than MvcA-Lpg2149, which is consistent with structural analysis.this study fully expounds the catalytic and regulatory mechanism of Mavc-mediated new ubiquitin modification, which not only helps to understand the diversity mechanism of the game between pathogens and hosts, but also provides ideas for the design of new anti-bacterial therapies.
researcher of the Institute of Biophysics is the communication author of this paper, and Wang Yong, associate researcher of his research group, and Zhang Qi, ph.D. student, are the co-authors of this paper. Wang Xinlu, a researcher at the Institute of Biophysics and an associate researcher in his research group, guided and carried out cell-related experiments. (Source: China Science Journal Gan Xiao Zhanqi)
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