Comparison of the catalytic and immunological properties of beta-glucosidases from three strains of Saccharomyces lactis

beta-Glucosidase from Saccharomyces lactis strains Y-123 (B(h)), Y-14 (B(m)), and Y-1057A (B(1)) was partially purified. The pH optima, Michaelis constants, and activation energies were determined for the hydrolysis of p-nitro-phenyl-beta-d-glucoside by each of the enzymes. Differences among these constants were not enough to account for the low specific activity of beta-glucosidase in strains Y-14 and Y-1057A. Enzyme-inhibitor constants were measured for a series of alkyl and aryl glucosides. In general, the three enzymes are arylglucosidases. Tris(hydroxymethyl)-aminomethane inhibited all three enzymes in an uncompetitive fashion. The inhibition was antagonized by Mg(++). An antiserum was prepared to the highly purified (200-fold) beta-glucosidase from strain Y-123. The nature and degree of cross-reaction between the three beta-glucosidases was investigated by double diffusion in agar and neutralization tests. Spur formation in the immunodiffusion tests and similar equivalence points in the neutralization tests indicated a strong degree of cross-reaction between the three enzymes. The ratio of enzyme activity to antigenicity was used to compare the relative molecular activity of beta-glucosidase in the three strains. Each strain produced the same amount of beta-glucosidase per milligram of cell protein. The results are consistent either with a lower turnover number for the beta-glucosidase in strains Y-14 and Y-1057A or with the production of beta-glucosidase with a "normal" turnover number and enough cross-reacting material to effectively reduce the specific activity to the observed levels.     

Similar hypotensive responses to resistance exercise with and without blood flow restriction

Low intensity resistance exercise (RE) with blood flow restriction (BFR) has gained attention in the literature due to the beneficial effects on functional and morphological variables, similar to those observed during traditional RE without BFR, while the effects of BFR on post-exercise hypotension remain unclear. The aim of the present study was to compare the blood pressure (BP) response of trained normotensive individuals to RE with and without BFR. In this cross-over randomized trial, eight male subjects (23.8 ± 4 years, 74 ± 3 kg, 174 ± 4 cm) completed two exercise protocols: traditional RE (3 x 10 repetitions at 70% one-repetition maximum [1-RM]) and low intensity RE (3 x 15 repetitions at 20% 1-RM) with BFR. Blood pressure measurements were performed after 15 min of seated rest (0), immediately after and 10 min, 20 min, 30 min, 40 min, 50 min and 60 min after the experimental sessions. Similar hypotensive effects for systolic BP (SBP) were observed for both protocols (P < 0.05) after exercise, with no differences between groups (P > 0.05) and no statistically significant difference for diastolic BP (P > 0.05). These results suggest that in normotensive trained individuals, both traditional RE and RE with BFR induce hypotension for SBP, which is important to prevent cardiovascular disturbances.     

Long-term treadmill exercise improves spatial memory of male APPswe/PS1dE9 mice by regulation of BDNF expression and microglia activation

Increasing evidence suggests that physical activity could delay or attenuate the symptoms of Alzheimer''s disease (AD). But the underlying mechanisms are still not fully understood. To investigate the effect of long-term treadmill exercise on the spatial memory of AD mice and the possible role of β-amyloid, brain-derived neurotrophic factor (BDNF) and microglia in the effect, male APPswe/PS1dE9 AD mice aged 4 months were subjected to treadmill exercise for 5 months with 6 sessions per week and gradually increased load. A Morris water maze was used to evaluate the spatial memory. Expression levels of β-amyloid, BDNF and Iba-1 (a microglia marker) in brain tissue were detected by immunohistochemistry. Sedentary AD mice and wildtype C57BL/6J mice served as controls. The results showed that 5-month treadmill exercise significantly decreased the escape latencies (P < 0.01 on the 4th day) and improved the spatial memory of the AD mice in the water maze test. Meanwhile, treadmill exercise significantly increased the number of BDNF-positive cells and decreased the ratios of activated microglia in both the cerebral cortex and the hippocampus. However, treadmill exercise did not significantly alleviate the accumulation of β-amyloid in either the cerebral cortex or the hippocampus of the AD mice (P > 0.05). The study suggested that long-term treadmill exercise could improve the spatial memory of the male APPswe/PS1dE9 AD mice. The increase in BDNF-positive cells and decrease in activated microglia might underpin the beneficial effect.     

Further characterization of isolated presumptive euchromatin fractions of mouse hepatoma cells utilizing a cDNA probe complementary to the homologous poly(A)+ mRNA

Poly(A)+ mRNA from mouse hepatoma ascites cell cytoplasm is characterized by three frequency classes: an abundant frequency class of a limited number of different nucleotide sequences, a less abundant frequency class of a larger number of different nucleotide sequences, and a rare frequency class containing a high number of different nucleotide sequences. [3H]cDNA synthesized on this poly(A)+ mRNA template hybridizes with some of the DNAs of the putative transcribable euchromatin fraction at a significantly faster rate than with total DNA if residual contaminating RNA is not removed. Following NaOH incubation to remove such RNA, the cDNA probe hybridized with essentially the same rate to the euchromatin fractions and total DNA. Nick translation of the nuclease-sensitive sequences of chromatin demonstrated that, even with limited nuclease digestion, the excised sequences rapidly converted to small oligonucleotides. The nick-translatable, small chromatin segments showed no enrichment for transcribable sequences. Chromatin segments, which distribute to the 50S-70S glycerol gradient fractions and which satisfy several of the presumptive criteria for enrichment for transcribable sequences, therefore show no enrichment for sequences complementary to the cDNA for poly(A)+ mRNA.     

Release of euchromatic segments by MgCa-dependent autodigestion of chromatin

Mouse TLT hepatoma chromatin was incubated in the presence of added MgCl₂ and CaCl₂ but in the absence of added exogenous nuclease. Fraction-ation of such autodigested chromatin by glycerol density gradient centrifugation resulted in a heterochromatin-enriched pellet and euchromatin-enriched non-pelletable fractions. This was determined by satellite DNA content analyses and analysis of nascent RNA distribution. Electron microscope examination of the chromatin revealed it to consist of a series of connected spherical particles (nucleosomes) having a diameter of 370 ± 70Å. This native structure of chromatin was not damaged by the autodigestion process.