Structural insights into the molecular mechanism underlying Sirt5-catalyzed desuccinylation of histone peptides.

Histone modification is a ubiquitous regulatory mechanism involved in a variety of biological processes, including gene expression, DNA damage repair, cell differentiation, and ontogenesis. Succinylation sites on histones have been identified and may have functional consequences. Here, we demonstrate that human sirtuin 5 (Sirt5) catalyzes the sequence-selective desuccinylation of numerous histone succinyl sites. Structural studies of Sirt5 in complex with four succinyl peptides indicate an essential role for the conserved main chain hydrogen bonds formed by the succinyl lysine (0), +1, and +3 sites for substrate-enzyme recognition. Furthermore, biochemical assays reveal that the proline residue at the +1 site of the histone succinylation substrate is unfavorable for Sirt5 interaction. Our findings illustrate the molecular mechanism underlying the sequence-selective desuccinylase activity of Sirt5 and provide insights for further studies of the biological functions associated with histone succinylation and Sirt5.

Structural insights into a novel functional dimer of Staphylococcus aureus RNase HII.

RNase HII exists ubiquitously in organisms and functions as a monomer in prokaryotes. We determined the crystal structure of Staphylococcus aureus RNase HII (Sa-RNase HII), which displays a novel dimer conformation, with the active site of each monomer covered by the other one. Both small-angle X-ray scattering and gel-filtration analysis confirmed that Sa-RNase HII exists as a homodimer in solution. Enzymatic analysis revealed that the "self-inhibited" dimeric form is catalytically active. Furthermore, continuous-wave electron paramagnetic resonance experiments clarified that the Sa-RNase HII dimer undergoes a large conformational change upon substrate binding, but remains a dimer to catalyze the reaction. Our structural and biochemical studies identified a novel functional dimer of Sa-RNase HII with distinct regulation mechanism for its catalytic activity.

Staphylococcus aureus SdrE captures complement factor H's C-terminus via a novel 'close, dock, lock and latch' mechanism for complement evasion.

Complement factor H (CFH) is a soluble complement regulatory protein essential for the down-regulation of the alternative pathway on interaction with specific markers on the host cell surface. It recognizes the complement component 3b (C3b) and 3d (C3d) fragments in addition to self cell markers (i.e. glycosaminoglycans, sialic acid) to distinguish host cells that deserve protection from pathogens that should be eliminated. The Staphylococcus aureus surface protein serine-aspartate repeat protein E (SdrE) was previously reported to bind human CFH as an immune-evasion tactic. However, the molecular mechanism underlying SdrE-CFH-mediated immune evasion remains unknown. In the present study, we identified a novel region at CFH's C-terminus (CFH1206-1226), which binds SdrE N2 and N3 domains (SdrEN2N3) with high affinity, and determined the crystal structures of apo-SdrEN2N3 and the SdrEN2N3-CFH1206-1226 complex. Comparison of the structure of the CFH-SdrE complex with other CFH structures reveals that CFH's C-terminal tail flips from the main body to insert into the ligand-binding groove of SdrE. In addition, SdrEN2N3 adopts a 'close' state in the absence of CFH, which undergoes a large conformational change on CFH binding, suggesting a novel 'close, dock, lock and latch' (CDLL) mechanism for SdrE to recognize its ligand. Our findings imply that SdrE functions as a 'clamp' to capture CFH's C-terminal tail via a unique CDLL mechanism and sequesters CFH on the surface of S. aureus for complement evasion.

Structure insights into the molecular mechanism of the interaction between UHRF2 and PCNA.

UHRF2 (Ubiquitin-like with PHD and ring finger domains 2) is an E3 ubiquitin ligase that plays important roles in DNA methylation, histone modifications and cell cycle regulation by interacting with multiple epigenetic or cell-cycle related proteins. Previous studied have identified PCNA (Proliferating cell nuclear antigen) as an interacting partner of UHRF2 by using the antibody microarray. However, the molecular mechanism and the function of UHRF2-PCNA interaction remains unclear. Here, we report the complex structure of PCNA and the peptide (784NEILQTLLDLFFPGYSK800) derived from UHRF2 that contains a PIP box. Structural analysis combined with mutagenesis experiments provide the molecular basis for the recognition of UHRF2 by PCNA via PIP-box.

Structure of the JmjC domain-containing protein NO66 complexed with ribosomal protein Rpl8.

The JmjC domain-containing proteins belong to a large family of oxygenases possessing distinct substrate specificities which are involved in the regulation of different biological processes, such as gene transcription, RNA processing and translation. Nucleolar protein 66 (NO66) is a JmjC domain-containing protein which has been reported to be a histone demethylase and a ribosome protein 8 (Rpl8) hydroxylase. The present biochemical study confirmed the hydroxylase activity of NO66 and showed that oligomerization is required for NO66 to efficiently catalyze the hydroxylation of Rpl8. The structures of NO66(176-C) complexed with Rpl8(204-224) in a tetrameric form and of the mutant protein M2 in a dimeric form were solved. Based on the results of structural and biochemical analyses, the consensus sequence motif NHXH recognized by NO66 was confirmed. Several potential substrates of NO66 were found by a BLAST search according to the consensus sequence motif. When binding to substrate, the relative positions of each subunit in the NO66 tetramer shift. Oligomerization may facilitate the motion of each subunit in the NO66 tetramer and affect the catalytic activity.

The modified outer membrane protein Amuc_1100 of Akkermansia muciniphila improves chronic stress-induced anxiety and depression-like behavior in mice.

Akkermansia muciniphila is a next-generation probiotic. The interaction between outer membrane protein Amuc_1100 of A. muciniphila and toll-like receptor 2 (TLR2) in intestinal epithelial cells influences the level of intestinal 5-hydroxytryptamine (5-HT). Amuc_1100Δ80 is a truncated form of Amuc_1100 lacking the first 80 N-terminal amino acids and has a higher affinity for TLR2 than the wild-type protein. Here, we report that Amuc_1100Δ80 could significantly reduce anxiety and depression-like behavior of mice when they were exposed to chronic unpredictable mild stress (CUMS). The experimental results of the rat insulinoma cell line RIN-14B showed that Amuc_1100Δ80 also induced a significantly higher upregulation of tryptophan hydroxylase 1 (Tph1), a rate-limiting enzyme of intestinal 5-HT synthesis. The imbalance of the gut microflora could be diminished when CUMS mice were fed with Amuc_1100Δ80. These results reveal that Amuc_1100Δ80 could affect the 5-HT level and the downstream 5-HTR1A-CREB-BDNF signal pathway via interacting with TLR2 and by altering the gut microbial composition. In parallel, the downregulation exerted by Amuc_1100Δ80 on the inflammation and hyperactivated HPA axis was closely related to the improvement of depression-like symptoms in CUMS mice. This study not only provides new insights into the antidepressant effect of A. muciniphila and its outer membrane protein Amuc_1100 but also identifies new potential targets and pathways in the gut for future research and the development of antidepressant drugs.

Structural insights into the intermolecular interaction of the adhesin SdrC in the pathogenicity of Staphylococcus aureus.

Staphylococcus aureus is an opportunistic disease-causing pathogen that is widely found in the community and on medical equipment. A series of virulence factors secreted by S. aureus can trigger severe diseases such as sepsis, endocarditis and toxic shock, and thus have a great impact on human health. The transformation of S. aureus from a colonization state to a pathogenic state during its life cycle is intimately associated with the initiation of bacterial aggregation and biofilm accumulation. SdrC, an S. aureus surface protein, can act as an adhesin to promote cell attachment and aggregation by an unknown mechanism. Here, structural studies demonstrate that SdrC forms a unique dimer through intermolecular interaction. It is proposed that the dimerization of SdrC enhances the efficiency of bacteria-host attachment and therefore contributes to the pathogenicity of S. aureus.