Characterization of Nocardithiocin Derivatives Produced by Amino Acid Substitution of Precursor Peptide notG

International Journal of Peptide Research and Therapeutics - Nocardithiocin is a thiopeptide compound produced by the pathogenic actinomycete Nocardia pseudobrasiliensis that displays activity...     

Bovine lactoferrin promotes energy expenditure via the cAMP-PKA signaling pathway in human reprogrammed brown adipocytes

Lactoferrin (LF) is a multifunctional protein in mammalian milk. We previously reported that enteric-coated bovine LF reduced the visceral fat in a double-blind clinical study. We further demonstrated that bovine LF (bLF) inhibited adipogenesis and promoted lipolysis in white adipocytes, but the effect of bLF on brown adipocytes has not been clarified. In this study, we investigated the effects of bLF on energy expenditure and cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signaling pathway using human reprogrammed brown adipocytes generated by gene transduction. bLF at concentrations of ≥?100 μg/mL significantly increased uncoupling protein 1 (UCP1) mRNA levels, with the maximum value observed 4 h after bLF addition. At the same time point, bLF stimulation also significantly increased oxygen consumption. Signaling pathway analysis revealed rapid increases of intracellular cAMP and cAMP response element-binding protein (CREB) phosphorylation levels beginning 5 min after bLF addition. The mRNA levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) were also significantly increased after 1 h of bLF stimulation. H-89, a specific PKA inhibitor, abrogated bLF-induced UCP1 gene expression. Moreover, receptor-associated protein (Rap), an antagonist of low-density lipoprotein receptor-related protein 1 (LRP1), significantly reduced bLF-induced UCP1 gene expression in a dose-dependent manner. These results suggest that bLF promotes UCP1 gene expression in brown adipocytes through the cAMP-PKA signaling pathway via the LRP1 receptor, leading to increased energy expenditure.     

Chaperone-mediated reduction of RepA dimerization is associated with RepA conformational change

RepA, the initiator protein of plasmid P1, binds to multiple sites (iterons) in the origin. The binding normally requires participation of chaperones, DnaJ, DnaK and GrpE. When purified, RepA appears dimeric and is inactive in iteron binding. On reaction with chaperones, a species active in iteron binding is formed and found to be monomeric. To test whether the chaperones can reduce dimerization, RepA was used to replace the dimerization domain of the λ repressor. The hybrid protein repressed the λ operator efficiently, indicating that RepA can dimerize in vivo . A further increase in repressor activity was seen in dnaJ mutant cells. These results are consistent with a chaperone-mediated reduction of RepA dimerization. We also found that RepA mutants defective in dimerization still depend on DnaJ for iteron binding. Conversely, RepA mutants that no longer require chaperones for iteron binding remain dimerization proficient. These results indicate that the chaperone dependence of RepA activity is not solely owing to RepA dimerization. Our results are most simply explained by a chaperone-mediated conformational change in RepA protomer that activates iteron binding. This conformational change also results in reduced RepA dimerization.     

Replication of mini-F plasmid in vitro promoted by purified E protein

Summary An in vitro system for replication of mini-F plasmid DNA was constructed. This system consists of an ammonium sulfate fraction II (Fuller et al. 1981) from Escherichia coli extract, exogeneously added purified E protein encoded by mini-F plasmid, and mini-F DNA in a closed circular form. Experiments with this system showed that the 217 bp DNA region which contains the A+T rich cluster and the four 19 bp direct repeats responsible for incB incompatibility is essential for mini-F DNA replication.     

Polyclonal B Cell Activation by Cell Wall Preparations of Gram-Positive Bacteria

The effects of polyclonal B cell activation (PBA) of cell walls and their cell wall fractions obtained from several kinds of gram-positive bacteria were studied using the anti-sheep red blood cell (SRBC) or anti-trinitrophenylated (TNP) SRBC plaque forming cell (PFC) responses of cultured spleen cells from Balb/c, athymic nu/nu, their littermates (nu/+), C3H/He (LPS-responder), C3H/HeJ (LPS-non-responder), (CBA/N × Balb/c) F1 male with an X-linked defect in B cell function and the F1 female mice. The cell walls of Staphylococcus epidermidis (ATCC 155), Lactobacillus plantarum (ATCC 8014), Micrococcus lysodeikticus (NCTC 2665), Mycobacterium rhodochrous (ATCC 184), Streptomyces gardneri (ATCC 23911) and Nocardia corynebacteriodes (ATCC 14898) had the ability to induce polyclonal B cell responses in the spleen cells of Balb/c, nu/nu, nu/+, C3H/He and C3H/HeJ mice. The cell wall fractions prepared by enzymatic digestion from the cell walls of S. epidermidis, S. gardneri or N. corynebacteriodes were also capable of inducing polyclonal B cell responses. The responses of spleen cells from (CBA/N × Balb/c) F1 male mice to these active preparations, except the cell walls of M. rhodochrous, were much lower than those of the F1 female mice. These findings indicate that the majority of the cell wall preparations lacks PBA ability for spleen cells with the CBA/N defect, except for the cell walls of M. rhodochrous which possess this ability. The PBA-ability of synthetic peptidoglycan, muramyl dipeptide (N-acetylmuramyl-L -alanyl-D -isoglutamine, MDP), was also examined, and a similar activity was observed in MDP.     

Anticancer saponin OSW-1 is a novel class of selective Golgi stress inducer

OSW-1 is a plant-derived natural product proposed to selectively kill cancer cells by binding to members of the oxysterol binding protein family, thereby disrupting lipid/sterol homeostasis. However, how these protein-ligand interactions mediate cell death signaling has remained elusive. Here, we discovered that OSW-1 selectively activates the Golgi stress response leading to apoptosis, providing a mechanistic basis for the anticancer activity of OSW-1.