Notes

18S rRNA gene sequence-structure phylogeny with the Trypanosomatida (Kinetoplastea, Euglenozoa) together with special mention of the Trypanosoma.
Additionally, we used generalised estimating equations to identify trial characteristics associated with intervention reporting. RESULTS Trials in our sample reported 8.6/12 TIDieR checklist items, on average. The most under-reported items were those for describing the expertise of the interventionists and for describing the location of the intervention. CONCLUSION Improved outcome reporting and intervention reproducibility among RCTs are greatly needed in cardiovascular medicine. Clinicians and researchers should advocate for the ethical publication of complete, translatable and replicable clinical research results. © Author(s) (or their employer(s)) 2020. BAY2666605 No commercial re-use. See rights and permissions. Published by BMJ.The C-type lectin receptors (CLRs) form a family of pattern recognition receptors (PRRs) that recognize numerous pathogens, such as bacteria and fungi, and trigger innate immune responses. The extracellular carbohydrate recognition domain (CRD) of CLRs forms a globular structure that can coordinate a Ca2+ ion, allowing receptor interactions with sugar-containing ligands. Although well conserved, the CRD fold can also display differences that directly affect the specificity of the receptors for their ligands. Here, we report crystal structures at 1.8-2.3 Å resolutions of the CRD of murine dendritic cell-immunoactivating receptor (DCAR/Clec4b1), the only CLR that binds phosphoglycolipids such as acylated phosphatidyl-myo-inositol mannosides (AcPIMs) of mycobacteria. Using mutagenesis analysis, we identified critical residues, Ala136 and Gln198, on the surface surrounding the ligand-binding site of DCAR, as well as an atypical Ca2+-binding motif (Glu-Pro-Ser/EPS168-170). By chemically synthesizing a water-soluble ligand analog, inositol-monophosphate di-mannose (IPM2), we confirmed the direct interaction of DCAR with the polar moiety of AcPIMs by biolayer interferometry and co-crystallization approaches. We also observed a hydrophobic groove extending from the ligand-binding site that is in a suitable position to interact with the lipid portion of whole AcPIMs. These results suggest that the hydroxyl group-binding ability and hydrophobic groove of DCAR mediate its specific binding to pathogen-derived phosphoglycolipids such as mycobacterial AcPIMs. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.β-Mannanases from the glycoside hydrolase 26 (GH26) family are retaining hydrolases that are active on complex heteromannans and whose genes are abundant in rumen metagenomes and metatranscriptomes. These enzymes can exhibit distinct modes of substrate recognition and are often fused to carbohydrate-binding modules (CBMs), resulting in a molecular puzzle of mechanisms governing substrate preference and mode of action that has not yet been pieced together. In this study, we recovered a novel GH26 enzyme with a CBM35 module linked to its N-terminus (CrMan26) from a cattle rumen metatranscriptome. CrMan26 exhibited a preference for galactomannan as substrate and the crystal structure of the full-length protein at 1.85 Å resolution revealed a unique orientation of the ancillary domain relative to the catalytic interface, strategically positioning a surface aromatic cluster of the ancillary domain as an extension of the substrate-binding cleft, contributing to galactomannan preference. Moreover, systematic investigation of non-conserved residues in the catalytic interface unveiled that residues Tyr195 (-3 subsite) and Trp234 (-5 subsite) from distal negative subsites have key role in galactomannan preference. These results indicate a novel and complex mechanism for substrate recognition involving spatially remote motifs, distal negative subsites from the catalytic domain and a surface-associated aromatic cluster from the ancillary domain. These findings expand our molecular understanding of the mechanisms of substrate binding and recognition in the GH26 family and shed light on how some CBMs and their respective orientation can contribute to substrate preference. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Dynein light chain 8 (LC8) interacts with intrinsically disordered proteins (IDPs) and influences a wide range of biological processes. It is becoming apparent that among the numerous IDPs that interact with LC8, many contain multiple LC8-binding sites. Although it is established that LC8 forms parallel IDP duplexes with some partners, such as nucleoporin Nup159 and dynein intermediate chain, the molecular details of these interactions, and LC8's interactions with other diverse partners, remain largely uncharacterized. LC8 dimers could bind in either a paired "in-register" or in a heterogeneous "off-register" manner to any of the available sites on a multivalent partner. Here, using NMR chemical shift perturbation, analytical ultracentrifugation, and native electrospray ionization MS, we show that LC8 forms a stable "in-register" complex when bound to an IDP domain of the multivalent regulatory protein ATM/ATR-substrate CHK2-interacting zinc finger protein (ASCIZ). Using saturation transfer difference NMR, we demonstrate that at sub-stoichiometric LC8 concentrations, the IDP domain preferentially binds to one of the three LC8 recognition motifs, showing for the first time that the binding process is "in-register." Further, the dynamic behavior for the three sites and the size of the fully bound complex confirmed an "in-register" complex. Dynamics measurements also revealed that coupling between sites is dependent on the linker length separating these sites. These results identify linker length and motif specificity as drivers of "in-register" binding in the multivalent LC8/IDP complex assembly. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.Receptor-type protein tyrosine phosphatase α (RPTPα) is an important positive regulator of SRC kinase activation and a known promoter of cancer growth, fibrosis, and arthritis. The domain structure of RPTPs comprises an extracellular region, a transmembrane helix, and two tandem intracellular catalytic domains referred as D1 and D2. The D2 domain of RPTPs is believed to mostly play a regulatory function; however, no regulatory model has been established for RPTPα-D2 or other RPTP-D2 domains. Here, we solved the 1.8 Å resolution crystal structure of the cytoplasmic region of RPTPα, encompassing D1 and D2, trapped in a conformation that revealed a possible mechanism through which D2 can allosterically inhibit D1 activity. Using a D2-truncation RPTPα variant and mutational analysis of the D1/D2 interfaces, we show that D2 inhibits RPTPα phosphatase activity and identified a P405FTP408 motif in D1 that mediates the inhibitory effect of D2. Expression of the gain-of-function F406A/T407A RPTPα variant in HEK293T cells enhanced SRC activation, supporting the relevance of our proposed D2-mediated regulation mechanism in cell signaling.
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