Iron sulfate inhibits Limulus activity by induction of structural and qualitative changes in lipid A.

AIMS: The bacterial endotoxins test (BET) is a sensitive assay for measuring endotoxin levels in solution and uses the limulus amebocyte lysate (LAL) coagulation reaction. We sought to identify the mechanisms through which certain substances interfere with the interaction between LAL and bacterial lipopolysaccharide (LPS). METHODS AND RESULTS: Endotoxin lipid A was inactivated by the addition of iron sulfate, which acted on endotoxin directly and strongly inhibited LAL coagulation activity. Size-exclusion, anion-exchange and reverse-phase liquid chromatography/mass spectrometry were used to examine changes in inactivated lipid A in terms of complex formation, negative charge status, hydrophobic interaction and structure. Furthermore, we verified the involvement of the lipid A phosphoryl group in the interference of iron sulfate with lipid A-factor C binding activity. Iron sulfate-inactivated lipid A was cleaved at its glycosidic bond, resulting in loss of hydrophobic interactions and disruption of lipid A complexes without alteration of negative charge status and lipid A-factor C interaction. CONCLUSIONS: Lipid A cleavage was a direct result of interfering factors, including iron sulfate, which acted on endotoxin directly to disrupt lipid A complexes rather than interfering with LAL coagulation. SIGNIFICANCE AND IMPACT OF THE STUDY: Our data provide new insights into the mechanisms of lipid A activity.

The effect of short-term (10- and 15-min) running at self-selected intensity on mood alteration.

The primary purpose of this study was to contrast the effects of two short-duration exercises on mood changes. A secondary goal was to examine the relationship between pre-exercise and post-exercise mood states. The subjects were 15 healthy male graduate students. They were involved in a within-subject design in which each individual completed two trials of running on a treadmill, one trial for 10 minutes and the other for 15 minutes. The Mood Checklist Short-form 1 (MCL-S1) used in the present study represents the participants' mood states before, during, and after exercise. This questionnaire has three sub-scales measuring (a) pleasantness, (b) relaxation, and (c) anxiety. Participants ran on a treadmill for the assigned time at a self-selected intensity after being told to run at a rate that felt good and was not painful. ANOVA results showed that both of the short bouts of exercise affected the subjects' mood, because the main effect of time spent exercising was observed in all sub-scales of the MCL-S1 and there were no significant differences in trial-by-time interaction. In addition to these results, there was a significant correlation between the two trial lengths in the amount of pleasantness and the amount of anxiety felt at post-exercise. There were moderate differences in the effect size (ES) for pre- and post-exercise pleasantness and anxiety levels. These results revealed similar patterns of change. It seems reasonable to conclude, based on this study, that exercise between 10 and 15 minutes results in similar psychological benefits for the person exercising.

Application of chemical selective cleavage methods to analyze post-translational modification in proteins.

Three chemical specific cleavage reactions, one for the carboxyl side of aspartyl peptide bonds, one for the carboxyl side of asparaginyl peptide bonds and another for the amino side of seryl/threonyl peptide bonds have been recently established. Additionally, these reactions simultaneously react on several post-translationally modified groups in peptides or proteins. The modified groups cover the external modifications N-formyl, N-acetyl, N-pyroglutamyi residues and C-terminal-alpha amide, as well as the internal modifications such as O-acetyl serine, phosphorylated serine/tyrosine, sulfonylated tyrosine, glycosylated serine/threonine and glycosylated asparagine. These three cleavage reactions relate to key amino acids for modifications, deamidation for asparagine, phosphorylation and acetylation for serine, and glycosylation for asparagine, serine and threonine. The chemical reactions on these modifications change the peptide mapping pattern, and information from these reactions may contribute characterization and location of post-translational modified groups in the protein.