Tuberculosis, caused by Mycobacterium tuberculosis (Mtb), has threaten human health for thousands years. The chaperone trigger factor (TF) of Mtb (mtbTF), a ribosome-associated molecule, plays important roles in co-translational nascent chain folding and post-translational protein assembly. However, due to lack of structural information, the dynamic regulatory mechanism of mtbTF remains barely investigated. Herein we report the structural basis of the complex of TF and ribosomal protein S7 (mtbS7) from Mtb. The mtbTF-mtbS7 complex was obtained with high purity and homogeneity in vitro. MtbTF bound with mtbS7 in a Kd value of 1.433 μM, and formed a complex with mtbS7 at 1:2 M ratios as shown by isothermal titration calorimetry. In addition, the crystal structure of mtbS7 was solved to a resolution at 1.8 Å, which was composed of six α-helices and two β-strands. Moreover, the molecular envelopes of mtbTF and mtbTF-mtbS7 complex were built and consisted with these homologous structures by small-angle X-ray scattering method. Our current findings might provide structural basis for understanding the molecular mechanism of TF in protein folding and the regulation of ribosomal assembly in Mtb.

Phosphofructokinase B (PfkB) belongs to the ribokinase family, which uses the phosphorylated sugar as substrate, and catalyzes fructose-6-phosphate into fructose-1,6-diphosphate. However, the structural basis of Mycobacterium marinum PfkB is not clear. Here, we found that the PfkB protein was monomeric in solution, which was different from most enzymes in this family. The crystal structure of PfkB protein from M. marinum was solved at a resolution of 2.21 Å. The PfkB structure consists of two domains, a major three-layered α/β/α sandwich-like domain characteristic of the ribokinase-like superfamily, and a second domain composed of four-stranded β sheets. Structural comparison analysis suggested that residues G236, A237, G238, and D239 could be critical for ATP catalysis and substrate binding of PfkB. Our current work provides new insights into understanding the mechanism of the glycolysis in M. marinum.

DUX4 plays critical role in the molecular pathogenesis of the neuromuscular disorder facioscapulohumeral muscular dystrophy and acute lymphoblastic leukemia in humans. As a master transcription regulator, DUX4 can also bind the promoters and activate the transcription of hundreds ZGA-associated genes. Here we report on the structural and biochemical studies of DUX4 double homeodomains (DUX4-DH), representing the only structures contain both homeodomain 1 (HD1) and homeodomain 2 (HD2). HD1 and HD2 adopt classical homeobox fold; via the helix inserted into the major groove and the N-terminal extended loop inserted into the minor groove, HD1 and HD2 recognize the box1 (5'-TAA-3') and box2 (5'-TGA-3') nucleotides of the consensus sequence, respectively. Among the box1 and box2 linking nucleotides (CCTAA), the two adenine residues are reported to be highly conserved; however, they are not directly recognized by DUX4-DH in the structures. Besides different nucleotides, our ITC analysis indicated that DUX4-DH can also tolerate various changes in the linker length. Our studies not only revealed the basis for target DNA recognition by DUX4, but also advanced our understanding on multiple gene activation by DUX4.