Evaluation of Carotid Arterial Intima-Media Thickness (IMT) and Its Relation to Clinical Parameters in Japanese Children

The aim of this study was to evaluate the carotid arterial intima-media thickness (IMT) and its relation to clinical parameters in Japanese children. Fifty-two healthy children (39 boys and 13 girls), aged 6-14 years, were enrolled in this cross-sectional investigation study. IMT of the common carotid artery was determined using ultrasonography. We also investigated anthropometric parameters, blood pressure (BP), lifestyles and blood examinations. The mean value of IMT was 0.4±0.1mm, which was lower than the normal value (1.0mm) in adults. IMT was positively correlated with age (r＝0.340) and height (r＝0.346) in boys, while it was positively correlated with body mass index (BMI) (r＝0.584) and diastolic BP (DBP) (r＝0.563) in girls. In addition, IMT was associated with sleeping hours and hours of watching television (TV) by using stepwise regression analysis. In conclusion, IMT increased with aging, and it was linked to some clinical parameters of atherosclerosis and lifestyles in children. Therefore, this reference data will be helpful for future assessment of age-related change in Japanese children in clinical practice, and IMT might be a good predictor of atherosclerosis in Japanese children

Characterization of L-Arginine Oxidase Made from L-Glutamate Oxidase

　L‒Glutamate oxidase (LGOX) from Streptomyces sp. X‒119‒6 has strict substrate specificity toward L‒glutamate. Recently, we solved the X‒ray crystal structure of LGOX and this revealed that Arg305 in the active site is the key residue involved in substrate recognition. Therefore, we created 19 mutant enzymes of R305X‒LGOX by saturation mutagenesis. One of them R305D‒LGOX, Arg305 substituted with Asp exhibited oxidase activity for L‒Arg. Optimum pH of R305D‒LGOX mutant enzyme was pH 8.5. Interestingly, the activity of R305D‒LGOX toward L‒Arg was inhibited by phosphate. And furthermore, the substrate specificity of R305D‒LGOX was affected by using buffer. The results of inhibition analysis suggest, that phosphate is a competitive inhibitor of R305D‒LGOX when L‒Arg is used as substrate. Kinetic analysis of R305D‒LGOX showed that Km value and kcat value of R305D‒LGOX toward l-Arg were 0.68 mM and 6.7 s-1 respectively. In this study, we showed that R305D‒LGOX mutant enzyme is a novel l-arginine oxidase and useful for l-arginine biosensor

ATP-dependent reversible association of proteasomes with multiple protein components to form 26S complexes that degrade ubiquitinated proteins in human HL-60 cells

AbstractThe role of proteasomes in ubiquitin (Ub)-dependent protein degradation was studied by analyzing lysates of human promyelocytic leukemia HL-60 cells by glycerol density gradient centrifugation. High succinyl-Leu-Leu-Val-Tyr-4-methylcoumaryl-7-amide hydrolyzing activity was found in the 26S fraction, whereas the 20S fraction containing proteaomes had no activity. Addition of 0.05% sodium dodecylsulfate to the latter fraction, however, induced marked activity. The 26S, but not the 20S fraction catalyzed ATP-dependent degradation of [125I]lysozyme-Ub conjugate. Depletion from the lysate of ATP caused complete shift of the active 26S complex to the latent 20S form, whereas in the lysate prepared from ATP-depleted cells, ATP converted 20S proteasomes to 26S complexes. The immunoprecipitated 26S complexes were found to consist of proteasomes and 13–15 other proteins ranging in size from 35 to 110 kDa. We conclude that in the lysate, latent proteasomes undergo reversible, ATP-dependent association with multiple protein components to form 26S complexes that catalyze ATP-dependent degradation of Ub-protein conjugates

Cloning, sequencing and expression of VAT, a CDC48/p97 ATPase homologue from the archaeon Thermoplasma acidophilum

AbstractA member of the AAA family of Mg2+-ATPases from the archaeon Thermoplasma acidophilum has been cloned and expressed in Escherichia coli. The protein, VCP-like ATPase of Thermoplasma acidophilum (VAT), is a homologue of SAV from Sulfolobus acidocaldarius and CdcH of Halobacterium salinarium, and belongs to the CDC48/VCP/p97 subfamily. The deduced product of the vat gene is 745 residues long (Mr 83 000), which has an optimal Mg2+-ATPase activity at 70°C. Electron microscopy shows the purified protein to form single and double homo-hexameric rings. Although the symmetry is different, the appearance of the complexes formed of two rings resembles the 20S proteasome and Hsp60/GroEL.©1997 Federation of European Biochemical Societies