Anopheles mosquitoes are vectors of malaria, a potentially fatal blood disease affecting half billion humans worldwide. These blood-feeding insects include in their antihemostatic arsenal potent thrombin inhibitor, the flexible and cysteine-less anophelin. Here, we present thorough structure-and-function analysis inhibition by anophelin, including 2.3-Å crystal structure human thrombin·anophelin complex. Anophelin residues 32–61 well-defined electron density, completely occupying long cleft...

Micronuclei are a hallmark of cancer and several other human disorders. Recently, micronuclei were implicated in chromothripsis, series massive genomic rearrangements that may drive tumor evolution progression. Here, we show Aurora B kinase mediates surveillance mechanism integrates error correction during anaphase with spatial control nuclear envelope reassembly to prevent formation. Using high-resolution live-cell imaging non-cancer cells, uncover lagging chromosomes more frequent than...

Rab GTPases regulate discrete steps in vesicular transport pathways. Rabs require activation by specific guanine nucleotide exchange factors (GEFs) that stimulate the of GDP for GTP. Rab27a controls motility and regulated exocytosis secretory granules related organelles. In melanocytes, regulates peripheral mature melanosomes recruiting melanophilin myosin Va. Here, we studied melanocytes. We identify Rab3GEP, previously isolated as a GEF Rab3a, non-redundant GEF. Similar to Rab27a-deficient...

Rab geranylgeranyl transferase (RGGT) catalyzes the post-translational (GG) modification of (usually) two C-terminal cysteines in GTPases. Here we studied mechanism geranylgeranylation reaction by bisphosphonate analogs which one phosphonate group is replaced a carboxylate (phosphonocarboxylate, PC). The phosphonocarboxylates used were 3-PEHPC, was previously reported, and 2-hydroxy-3-imidazo[1,2-a]pyridin-3-yl-2-phosphonopropionic acid ((+)-3-IPEHPC), >25-fold more potent related compound...

Rhipicephalus (Boophilus) microplus is an ectoparasite responsible for important decrease in meat, milk and leather production, caused both by cattle blood loss the transmission of anaplasmosis babesiosis. R. a rich source serine protease inhibitors, including trypsin inhibitors BmTI-A BmTI-6, subtilisin inhibitor BmSI, recently described thrombin inhibitor, boophilin. Boophilin double Kunitz-type with unusual ability to form ternary complex second (non-thrombin) proteinase molecule. The...

Abstract Accurate chromosome segregation relies on microtubule end conversion, the ill-understood ability of kinetochores to transit from lateral attachment durable association with dynamic plus-ends. The molecular requirements for this conversion and underlying biophysical mechanisms are elusive. We reconstituted in vitro using two kinetochore components: plus end–directed kinesin CENP-E microtubule-binding Ndc80 complex, combined surface a microbead. primary role is ensure close proximity...

CLASPs are conserved microtubule plus-end–tracking proteins that suppress catastrophes and independently localize to kinetochores during mitosis. Thus, ideally positioned regulate kinetochore–microtubule dynamics required for chromosome segregation fidelity, but the underlying mechanism remains unknown. Here, we found human CLASP2 exists predominantly as a monomer in solution, it can self-associate through its C-terminal kinetochore-binding domain. Kinetochore localization was independent of...

The tremendous social and economic impact of thrombotic disorders, together with the considerable risks associated to currently available therapies, prompt for development more efficient safer anticoagulants. Novel peptide-based thrombin inhibitors were identified using in silico structure-based design further validated vitro. best candidate compounds contained both l- d-amino acids, general sequence d-Phe(P3)-Pro(P2)-d-Arg(P1)-P1′-CONH2. P1′ position was scanned d-isomers natural or...

Rab GTPases are important determinants of organelle identity and regulators vesicular transport pathways. Consequently, each occupies a highly specific subcellular localization. However, the precise mechanisms governing targeting remain unclear. Guanine nucleotide exchange factors (GEFs), putative membrane-resident effector binding have all been implicated as critical targeting. Here, we address these issues using Rab27a to melanosomes model system. regulates motility lysosome-related...

The cysteine-less peptidic anticoagulants madanin-1 and madanin-2 from the bush tick Haemaphysalis longicornis are founding members of MEROPS inhibitor family I53. It has been previously suggested that madanins exert their functional activity by competing with physiological substrates for binding to positively charged exosite I (fibrinogen-binding exosite) α-thrombin. We hereby demonstrate competitive inhibition α-thrombin or involves enzyme's active site. Moreover, blood coagulation factors...

ABSTRACT Rab GTPases are compartment-specific molecular switches that regulate intracellular vesicular transport in eukaryotes. GDP/GTP exchange factors (GEFs) control activation, and current models propose localised regulated GEF activity is important targeting Rabs to specific membranes. Here, we investigated the mechanism of function using Rab27a GEF, Rab3GEP (also known as MADD), melanocytes a model. We show Rab3GEP-deficient (melan-R3GKO) manifest partial disruption melanosome...

Post-translational cycles of α-tubulin detyrosination and tyrosination generate microtubule diversity, the cellular functions which remain largely unknown. Here we show that regulates kinetochore-microtubule attachments to ensure normal chromosome oscillations timely anaphase onset during mitosis. Remarkably, detyrosinated levels near kinetochore plus-ends depend on direction motion metaphase. Proteomic analyses unveil KNL-1/MIS12/NDC80 (KMN) network forms core microtubule-binding site at...

Abstract The fine regulation of kinetochore microtubule dynamics during mitosis ensures proper chromosome segregation by promoting error correction and spindle assembly checkpoint (SAC) satisfaction. CLASPs are widely conserved plus-end-tracking proteins that regulate throughout the cell cycle independently localize to kinetochores mitosis. Thus, ideally positioned dynamics, but underlying molecular mechanism remains unknown. Here we found human CLASP2 can dimerize through its C-terminal...

Introdução: A parada cardiorrespiratória é um eventocrítico para o paciente dialítico, que necessita de umaequipe enfermagem na hemodiálise apta a realizaro procedimento ressuscitação cardiopulmonar.Justifica-se nesse contexto educação permanenteem enfermagem, por meio da simulação in situ,pode contribuir com construção das competências emressuscitação cardiopulmonar.Objetivo: Verificar se diferentes periodicidades deformação utilizando situ, influenciamna ressuscitaçãocardiopulmonar no...

Summary Micronuclei are a hallmark of cancer and other human disorders have recently been implicated in chromothripsis, series massive genomic rearrangements that may drive tumor evolution progression. Here we show Aurora B kinase mediates surveillance mechanism integrates error correction during anaphase with spatial control nuclear envelope reformation to protect against micronuclei formation cell division. Using high-resolution live-cell imaging non-cancer cells found lagging chromosomes...

The serine protease thrombin plays a major role in thrombosis and haemostasis. This has driven interest inhibitors as potential antithrombotic drugs. Here, the crystallization preliminary crystallographic analysis of human α-­thrombin complex with three noncovalent peptide general sequence d-Phe-Pro-d-Arg-P1′-CONH2 are reported. crystals belonged to orthorhombic space group P212121 diffracted beyond 1.3 Å resolution.

Boophilin is a tight-binding thrombin inhibitor composed of two canonical Kunitz-type domains in tandem arrangement. Thrombin-bound boophilin can inhibit second trypsin-like serine proteinase, most likely through the reactive loop its N-terminal Kunitz domain. Here, crystallization and preliminary crystallographic analysis isolated domain reported. The crystals belonged to orthorhombic space group P2(1)2(1)2(1) diffracted beyond 1.8 Å resolution using sealed-tube home source 0.87 at...