C-terminal Tail of β-Tubulin and its Role in the Alterations of Dynein Binding Mode

Dynein is a cytoskeletal molecular motor protein that moves along the microtubule (MT) and transports various cellular cargos during its movement. Using standard Molecular Dynamics (MD) simulation, Principle Component Analysis (PCA), and Normal Mode Analysis (NMA) methods, this investigation studied large-scale movements and local interactions of dynein’s Microtubule Binding Domain (MTBD) when bound to tubulin heterodimer subunits. Examination of the interactions between the MTBD segments, and their adjustments in terms of intra- and intermolecular distances at the interfacial area with tubulin heterodimer, particularly at α-H16, β-H18 and β-tubulin C-terminal tail (CTT), was the main focus of this study. The specific intramolecular interactions, electrostatic forces and the salt-bridge residue pairs were shown to be the dominating factors in orchestrating movements of the MTBD and MT interfacial segments in the dynein’s low-high affinity binding modes. Important interactions included β-Glu447 and β-Glu449 (CTT) with Arg3469 (MTBD-H6), Lys3472 (MTBD-H6-H7 loop) and Lys3479 (MTBD-H7); β-Glu449 with Lys3384 (MTBD-H8), Lys3386 and His3387 (MTBD-H1). The structural and precise position, orientation, and functional effects of the CTTs on the MT-MTBD, within reasonable cut-off distance for non-bonding interactions and under physiological conditions, are unavailable from the previous studies. The absence of the residues in the highly flexible MT-CTTs in the experimentally solved structures is perhaps in some cases due to insufficient data from density maps, but these segments are crucial in protein binding. The presented work contributes to the information useful for the MT-MTBD structure refinement

Full-length structural model of RET3 and SEC21 in COPI: identification of binding sites on the appendage for accessory protein recruitment motifs

COPI, a 600 kD heptameric complex (consisting of subunits α, β, γ, δ, ε, ζ, and β′) “coatomer,” assembles non-clathrin-coated vesicles and is responsible for intra-Golgi and Golgi-to-ER protein trafficking. Here, we report the three-dimensional structures of the entire sequences of yeast Sec21 (γ-COPI mammalian ortholog), yeast Ret3 (ζ-COPI mammalian ortholog), and the results of successive molecular dynamics investigations of the subunits and assembly based on a protein–protein docking experiment. The three-dimensional structures of the subunits in their complexes indicate the residues of the two subunits that impact on assembly, the conformations of Ret3 and Sec21, and their binding orientations in the complexed state. The structure of the appendage domain of Sec21, with its two subdomains—the platform and the β-sandwich, was investigated to explore its capacity to bind to accessory protein recruitment motifs. Our study shows that a binding site on the platform is capable of binding the Eps15 DPF and epsin DPW2 peptides, whereas the second site on the platform and the site on the β-sandwich subdomain were found to selectively bind to the amphiphysin FXDXF and epsin DPW1 peptides, respectively. Identifying the regions of both the platform and sandwich subdomains involved in binding each peptide motif clarifies the mechanism through which the appendage domain of Sec21 engages with the accessory proteins during the trafficking process of non-clathrin-coated vesicles

Cholinesterases: Structure, Role, and Inhibition

Acetilkolinesteraza (AChE; E.C. 3.1.1.7) i butirilkolinesteraza (BChE; E.C. 3.1.1.8) enzimi su koji se zbog svoje uloge u organizmu intenzivno istražuju unutar područja biomedicine i toksikologije. Iako strukturno homologni, ovi enzimi razlikuju se prema katalitičkoj aktivnosti, odnosno specifi čnosti prema supstratima koje mogu hidrolizirati te selektivnosti za vezanje mnogih liganada. U ovom radu dan je pregled dosadašnjih istraživanja kolinesteraza i njihovih interakcija s ligandima i inhibitorima te su izdvojene aminokiseline aktivnog mjesta koje sudjeluju u tim interakcijama.Enzymes acetylcholinesterase (AChE; E.C. 3.1.1.7) and butyrylcholinesterase (BChE; E.C. 3.1.1.8) have intensively been investigated in biomedicine and toxicology due to important role in organisms. Even if structurally homologous, they differ in catalytic activity, specificity, for substrates, and selectivity in binding to many ligands. This paper compiles the results of research on cholinesterases and their interactions with ligands and inhibitors, and identifies amino acids of active sites involved in these interactions

The importance of lipid peroxidation and iron for the ischaemia-reperfusion injury of kidneys

All over the world, scientists are studying the possible role of oxygen free radicals (OFR) in numerous pathological conditions, including ischaemia- reperfusion (I/R). The final step in the radical damage is lipid peroxidation. The radical damage is considerably increased in the presence of iron.Aims of the present study:1. To examine the effects of pre-treatment with desferrioxamine or H290/51 on the production of radicals at reperfusion after ischaemia.2. To examine the effects of pre-treatment with desferrioxamine or H290/51 on the kidney function and survival after ischaemia-reperfusion.3. To study the effects of ischaemia and reperfusion on lipid peroxidation as reflected in TBARS production in vivo and in vitro and to elucidate the effects of pre-treatment with H290/51 in this respect.4. To study the effect of ischaemia and reperfusion on mitochondrial bioenergetics and intracellular pH measured with volume-selective 31P NMR spectroscopy (OSIRIS) and to elucidate the effects of pretreat-ment with H290/51 in these respects. Methods: New Zealand white rabbits and Sprague-Dawley rats were used. In the ESR experiments, OXANOH was used as spin trap. In the TBARS experiment, the kidneys were sliced after ischaemia and put into Krebs solution and reoxygenated in a 37°C waterbath. Then the slices were chopped and frozen for later analyses of TBARS. In the NMR experiments, the rabbit was placed in a magnetic tube during the experiments and could therefore not be touched. An experimental model in which the blood supply to the renal artery could be clamped at a distance by means of a Fogarty-catheter was developed. Results: In the ESR studies, pre-treatment with desferrioxamine reduced the production of radicals to 30% compared with pre-ischaemic values, but pre-treatment with H290/51 completely abolished the increase in the production of radicals seen after ischaemia. Pre-treatment with desferrioxamine improved glomerular function, but had no effect on tubular function after I/R. Pre-treatment with H290/51 improved both glomerular and tubular functions after I/R. Lipid peroxidation as reflected in TBARS-production was reduced after pre-treatment with H290/51 to near pre-ischaemic values. Mitochondrial function after I/R, as reflected in ATP and pH increase, was greatly improved after pre-treatment with H290/51. Conclusions: Desferrioxamine and H290/51 diminish the production of radicals and protect renal function after I/R. H290/51 diminishes lipid peroxidation, as reflected in TBARS production, and protects the mitochondrial function after I/R