Partially Folded Structure of Flavin Adenine Dinucleotide-depleted Ferredoxin-NADP+ Reductase with Residual NADP+ Binding Domain

This research was originally published in the Journal of Biological Chemistry. Masahiro Maeda, Daizo Hamada, Masaru Hoshino, Yayoi Onda, Toshiharu Hase and Yuji Goto. Partially Folded Structure of Flavin Adenine Dinucleotide-depleted Ferredoxin-NADP+ Reductase with Residual NADP+ Binding Domain. J. Biol. Chem. 2002; 277, 17101-17107. © the American Society for Biochemistry and Molecular Biolog

Direct Observation of Amyloid Fibril Growth Monitored by Thioflavin T Fluorescence

This research was originally published in the Journal of Biological Chemistry. Tadato Ban, Daizo Hamada, Kazuhiro Hasegawa, Hironobu Naiki and Yuji Goto. Direct Observation of Amyloid Fibril Growth Monitored by Thioflavin T Fluorescence. J. Biol. Chem. 2003; 278, 16462-16465. © the American Society for Biochemistry and Molecular Biolog

High helicity of peptide fragments corresponding to β-strand regions of β-lactoglobulin observed by 2D-NMR spectroscopy

Backgound: Whereas protein fragments, when they are structured, adopt conformations similar to that found in the native state, the high helical propensity of β-lactoglobulin, a predominantly β-sheet protein, suggested that the fragments of β-lactoglobulin can assume the non-native helical conformation. In order to assess this possibility, we synthesized four 17–18-residue peptides corresponding to three β-strand regions and one helical region (as a control) of β-lactoglobulin and examined their conformation.ResultsWe observed residual helicities of up to 17% in water, by far-UV CD, for all four peptide fragments. The helices could be significantly stabilized by the addition of TFE, and the NMR analyses in a mixture of 50% water/TFE indicated that helical structures are formed in the central region whereas both termini are frayed. Thus, the very same residues that form strands in the native β-lactoglobulin showed high helical preferences.ConclusionThese results stand out from the current general view that peptide fragments isolated from proteins either are unfolded or adopt native-like secondary structures. The implications of the results in the mechanism of protein folding and in designing proteins and peptides are significant

N-Terminal Phosphorylation of HP1α Promotes Its Chromatin Binding ▿

The phosphorylation of heterochromatin protein 1 (HP1) has been previously described in studies of mammals, but the biological implications of this modification remain largely elusive. Here, we show that the N-terminal phosphorylation of HP1α plays a central role in its targeting to chromatin. Recombinant HP1α prepared from mammalian cultured cells exhibited a stronger binding affinity for K9-methylated histone H3 (H3K9me) than that produced in Escherichia coli. Biochemical analyses revealed that HP1α was multiply phosphorylated at N-terminal serine residues (S11-14) in human and mouse cells and that this phosphorylation enhanced HP1α's affinity for H3K9me. Importantly, the N-terminal phosphorylation appeared to facilitate the initial binding of HP1α to H3K9me by mediating the interaction between HP1α and a part of the H3 tail that was distinct from the methylated K9. Unphosphorylatable mutant HP1α exhibited severe heterochromatin localization defects in vivo, and its prolonged expression led to increased chromosomal instability. Our results suggest that HP1α's N-terminal phosphorylation is essential for its proper targeting to heterochromatin and that its binding to the methylated histone tail is achieved by the cooperative action of the chromodomain and neighboring posttranslational modifications

Binding of Islet Amyloid Polypeptide to Supported Lipid Bilayers and Amyloid Aggregation at the Membranes

Amyloid deposition of human islet amyloid polypeptide (hIAPP) in the islets of Langerhans is closely associated with the pathogenesis of type II diabetes mellitus. Despite substantial evidence linking amyloidogenic hIAPP to loss of β-cell mass and decreased pancreatic function, the molecular mechanism of hIAPP cytotoxicity is poorly understood. We here investigated the binding of hIAPP and nonamyloidogenic rat IAPP to substrate-supported planar bilayers and examined the membrane-mediated amyloid aggregation. The membrane binding of IAPP in soluble and fibrillar states was characterized using quartz crystal microbalance with dissipation monitoring, revealing significant differences in the binding abilities among different species and conformational states of IAPP. Patterned model membranes composed of polymerized and fluid lipid bilayer domains were used to microscopically observe the amyloid aggregation of hIAPP in its membrane-bound state. The results have important implications for lipid-mediated aggregation following the penetration of hIAPP into fluid membranes. Using the fluorescence recovery after photobleaching method, we show that the processes of membrane binding and subsequent amyloid aggregation are accompanied by substantial changes in membrane fluidity and morphology. Additionally, we show that the fibrillar hIAPP has a potential ability to perturb the membrane structure in experiments of the fibril-mediated aggregation of lipid vesicles. The results obtained in this study using model membranes reveal that membrane-bound hIAPP species display a pronounced membrane perturbation ability and suggest the potential involvement of the oligomeic forms of hAPP in membrane dysfunction

Diagnostics of the Performance Quality in Top Level Players

Název: Diagnostika herního výkonu u hráčů elitní výkonnosti Diagnostic ofthe duality performance by players ofelite performance level Cíle práce: A) Odhalit "chybná řešení" v obranné fázi u mužstva dospělých hráčů elitní výkonnosti. B) Kategorizovat zjištěné poznatky a navrhnout opatření pro tréninkový proces, která by přispěla ke zlepšení herního výkonu. Metoda: Použili jsme metody nepřímého pozorování pomocí videozáznamu.V šetření jsme využili kvantitativní a kvalitativní analýzy "chybných řešení" objevujících se v obranné fázi u vybraných utkání mužstva FC Chlesea v Lize mistrů. Výsledky: Analýza utkání odhalila opakující se skutečnosti v chybném herním výkonu mužstva při obranné fázi hry.Z této skutečnosti lze vyvodit požadavky, které jsou kladeny na současné hráče elitní výkonnosti. Klíčová slova: fotbal - chybovost - herní výkon - obranné systémySportovní hryFaculty of Physical Education and SportFakulta tělesné výchovy a sport