Lytic transglycosylase repertoire diversity enables intrinsic antibiotic resistance and daughter cell separation in Escherichia coli under acidic stress.

Peptidoglycan (PG) is an important architectural element that imparts physical toughness and rigidity to the bacterial envelope. It is also a dynamic structure that undergoes continuous turnover or autolysis. Escherichia coli possesses redundant PG degradation enzymes responsible for PG turnover; however, the advantage afforded by the existence of numerous PG degradation enzymes remains incompletely understood. In this study, we elucidated the physiological roles of MltE and MltC, members of the lytic transglycosylase (LTG) family that catalyze the cleavage of glycosidic bonds between disaccharide subunits within PG strands. MltE and MltC are acidic LTGs that exhibit increased enzymatic activity and protein levels under acidic pH conditions, respectively, and deletion of these two LTGs results in a pronounced growth defect at acidic pH. Furthermore, inactivation of these two LTGs induces increased susceptibility at acidic pH against various antibiotics, particularly vancomycin, which seems to be partially caused by elevated membrane permeability. Intriguingly, inactivation of these LTGs induces a chaining morphology, indicative of daughter cell separation defects, only under acidic pH conditions. Simultaneous deletion of PG amidases, known contributors to daughter cell separation, exacerbates the chaining phenotype at acidic pH. This suggests that the two LTGs may participate in the cleavage of glycan strands between daughter cells under acidic pH conditions. Collectively, our findings highlight the role of LTG repertoire diversity in facilitating bacterial survival and antibiotic resistance under stressful conditions.

Coordinated and Distinct Roles of Peptidoglycan Carboxypeptidases DacC and DacA in Cell Growth and Shape Maintenance under Stress Conditions.

Peptidoglycan (PG) is an essential bacterial architecture pivotal for shape maintenance and adaptation to osmotic stress. Although PG synthesis and modification are tightly regulated under harsh environmental stresses, few related mechanisms have been investigated. In this study, we aimed to investigate the coordinated and distinct roles of the PG dd-carboxypeptidases (DD-CPases) DacC and DacA in cell growth under alkaline and salt stresses and shape maintenance in Escherichia coli. We found that DacC is an alkaline DD-CPase, the enzyme activity and protein stability of which are significantly enhanced under alkaline stress. Both DacC and DacA were required for bacterial growth under alkaline stress, whereas only DacA was required for growth under salt stress. Under normal growth conditions, only DacA was necessary for cell shape maintenance, while under alkaline stress conditions, both DacA and DacC were necessary for cell shape maintenance, but their roles were distinct. Notably, all of these roles of DacC and DacA were independent of ld-transpeptidases, which are necessary for the formation of PG 3-3 cross-links and covalent bonds between PG and the outer membrane lipoprotein Lpp. Instead, DacC and DacA interacted with penicillin-binding proteins (PBPs)-dd-transpeptidases-mostly in a C-terminal domain-dependent manner, and these interactions were necessary for most of their roles. Collectively, our results demonstrate the coordinated and distinct novel roles of DD-CPases in bacterial growth and shape maintenance under stress conditions and provide novel insights into the cellular functions of DD-CPases associated with PBPs. IMPORTANCE Most bacteria have a peptidoglycan architecture for cell shape maintenance and protection against osmotic challenges. Peptidoglycan dd-carboxypeptidases control the amount of pentapeptide substrates, which are used in the formation of 4-3 cross-links by the peptidoglycan synthetic dd-transpeptidases, penicillin-binding proteins (PBPs). Seven dd-carboxypeptidases exist in Escherichia coli, but the physiological significance of their redundancy and their roles in peptidoglycan synthesis are poorly understood. Here, we showed that DacC is an alkaline dd-carboxypeptidase for which both protein stability and enzyme activity are significantly enhanced at high pH. Strikingly, dd-carboxypeptidases DacC and DacA physically interacted with PBPs, and these interactions were necessary for cell shape maintenance as well as growth under alkaline and salt stresses. Thus, cooperation between dd-carboxypeptidases and PBPs may allow E. coli to overcome various stresses and to maintain cell shape.

Distinct Amino Acid Availability-Dependent Regulatory Mechanisms of MepS and MepM Levels in Escherichia coli.

Peptidoglycan (PG) hydrolases play important roles in various aspects of bacterial physiology, including cytokinesis, PG synthesis, quality control of PG, PG recycling, and antibiotic resistance. However, the regulatory mechanisms of their expression are poorly understood. In this study, we have uncovered novel regulatory mechanisms of the protein levels of the synthetically lethal PG endopeptidases MepS and MepM, which are involved in PG synthesis. A mutant defective for both MepS and MepM was lethal in an amino acid-rich medium, whereas it exhibited almost normal growth in a minimal medium, suggesting the expendability of MepS and MepM in a minimal medium. Protein levels of MepS and MepM dramatically decreased in the minimal medium. Although MepM was revealed as a substrate of Prc, a periplasmic protease involved in the proteolysis of MepS, only the decrease in the MepS level in the minimal medium was affected by the prc depletion. Phenotypic and biochemical analyses showed that the presence of aromatic amino acids in the medium induced the accumulation of MepS, but not MepM, while the presence of glutamate increased the level of MepM, but not MepS. Together, these results demonstrate that the protein levels of the two major PG endopeptidases are regulated in an amino acid availability-dependent manner, but their molecular mechanisms and signaling are significantly distinct.

Development of a flickering action video based steady state visual evoked potential triggered brain computer interface-functional electrical stimulation for a rehabilitative action observation game.

BACKGROUND: This study focused on developing an upper limb rehabilitation program. In this regard, a steady state visual evoked potential (SSVEP) triggered brain computer interface (BCI)-functional electrical stimulation (FES) based action observation game featuring a flickering action video was designed. OBJECTIVE: In particular, the synergetic effect of the game was investigated by combining the action observation paradigm with BCI based FES. METHODS: The BCI-FES system was contrasted under two conditions: with flickering action video and flickering noise video. In this regard, 11 right-handed subjects aged between 22-27 years were recruited. The differences in brain activation in response to the two conditions were examined. RESULTS: The results indicate that T3 and P3 channels exhibited greater Mu suppression in 8-13 Hz for the action video than the noise video. Furthermore, T4, C4, and P4 channels indicated augmented high beta (21-30 Hz) for the action in contrast to the noise video. Finally, T4 indicated suppressed low beta (14-20 Hz) for the action video in contrast to the noise video. CONCLUSION: The flickering action video based BCI-FES system induced a more synergetic effect on cortical activation than the flickering noise based system.

Long-term effects of green tea ingestion on atherosclerotic biological markers in smokers.

OBJECTIVES: Smoking is a risk factor for coronary artery disease and triggers vascular injury by platelet aggregation and induces atherosclerosis through induction of oxidative stress. Green tea is known to have antioxidant capacity and anti-platelet activity. DESIGN AND METHODS: Twenty adult male smokers ingested 600 mL of green tea for 4 weeks. Their lipid profile, C-reactive protein (CRP), total antioxidant capacity, oxidized LDL, soluble VCAM-1, soluble ICAM-1, and soluble P-selectin were measured at baseline and 2 and 4 weeks after green tea ingestion. RESULTS: Plasma soluble P-selectin (sP-selectin) levels decreased significantly after 2 and 4 weeks of green tea ingestion compared with those before green tea ingestion (P < 0.001). Plasma concentrations of oxidized LDL decreased significantly after green tea ingestion (P < 0.05). CONCLUSIONS: The results of this study suggest the effect of green tea on sP-selectin and oxidized LDL.

Lack of association between oxidized LDL-cholesterol concentrations and haptoglobin phenotypes in healthy subjects.

BACKGROUND: The haptoglobin (Hp) 2-2 phenotype has been known to have less antioxidative activity than Hp 1-1 and Hp 2-1. However, few have reported the relationship between oxidative stress and Hp phenotypes by using oxidized LDL. The relationship between Hp phenotypes and oxidative stress was therefore investigated in healthy adults. METHODS: The serum Hp concentrations, albumin, uric acid, iron, L-ascorbic acid, total antioxidant status and oxidized LDL concentrations were measured and analysed in 200 healthy Korean men and women with Hp 2-1 and Hp 2-2 phenotypes. RESULTS: The serum concentrations of Hp were significantly higher in the Hp 2-1 compared with the Hp 2-2 phenotype. However, serum concentrations of total antioxidant status and oxidized LDL did not show any statistical differences in either sex. The L-ascorbic acid concentrations in serum showed lower values in the Hp 2-2 compared with the Hp 2-1 phenotype, for both sexes (P < 0.05). CONCLUSIONS: The concentration of L-ascorbic acid, the first line antioxidant, was lower in the Hp 2-2 than in the Hp 2-1 phenotype; this did not significantly affect the total antioxidant status, and there was no significant difference in oxidized LDL for either group in healthy adults.

Prevalence of carpal tunnel syndrome in meat and fish processing plants.

This study was carried out to survey the prevalence of carpal tunnel syndrome (CTS) in a high-risk job. The experimental group was composed of 69 workers sampled from meat and fish processing plants, who were using their upper extremities repetitively. The control group was composed of 28 workers sampled from managers, secretaries and keepers in the same companies. All the employees were examined for their work history, their symptoms, as well as physical examinations and a nerve conduction study (NCS). Fifty-one (73.9%) workers in the experimental group had findings compatible to the NIOSH definition of Work-Related CTS. The positive rates for Tinel and Phalen's signs were higher in the workers who had shorter working duration (<7 yr), but the positive rate in the NCS was higher in the workers who had longer working duration (>/=7 yr). The prevalence of CTS in a meat and fish processing plant was much higher. Therefore this job should be managed as a high-risk industry.

Low-density lipoprotein subclass and its correlating factors in diabetics.

OBJECTIVES: Small dense LDL, low density lipoprotein (LDL) particles with small size and high density, is regarded as a significant risk factor for cardiovascular diseases. Diabetes mellitus is one of the conditions accompanied by increased small dense LDL. We analyzed LDL subclass in type 2 diabetics and normal controls with LipoPrint LDL System to investigate the LDL heterogeneity in diabetics and factors affecting it. DESIGN AND METHODS: We selected 40 normal controls and 40 type 2 diabetics with fasting blood glucose level over 7.0 mmol/L and HbA1c level over 7%. LDL subclass was determined with LipoPrint LDL System. LipoPrint LDL System fractionates LDL into seven parts (LDL1-7) by size and LDL3 to LDL7 are defined as small-sized LDL. In addition we estimated 'the percent of small-sized LDL over whole LDL' and defined it as 'small-sized LDL proportion'. RESULTS: Mean small-sized LDL proportion was significantly higher in diabetics (23.4%) than in controls (11.8%) (p<0.001) and small-sized LDL proportion showed positive correlation with blood levels of glucose, HbA1c, total cholesterol, triglyceride, and oxidized LDL and negative correlation with HDL cholesterol level in univariate analysis (p<0.001). Of these parameters, triglyceride, HbA1c, oxidized LDL were statistically significant variables contributing to the small-sized LDL proportion in stepwise multiple regression analysis. CONCLUSIONS: We analyzed small-sized LDL proportion in type 2 diabetics and found that it was significantly increased in diabetics than control subjects and it was independently correlated with triglyceride, HbA1c, oxidized LDL in descending order, which are reflecting lipid metabolism, glycation, and oxidative stress, respectively.