Kinetic analysis of the interaction of tissue transglutaminase with a nonpeptidic slow-binding inhibitor

Tissue transglutaminase (TGase) is a Ca2+-dependent enzyme that catalyzes cross-linking of intracellular proteins through a mechanism that involves isopeptide bond formation between Gln and Lys residues and is allosterically regulated by GTP. TGase is thought to play a pathogenic role in neurodegenerative diseases by promoting aggregation of disease-specific proteins that accumulate as part of these disorders. Given the role that TGase plays in neurodegenerative disorders, we initiated a research program to discover inhibitors of this enzyme that might ultimately be developed into therapeutic agents. To identify such inhibitors, we screened 110,000 druglike compounds for their ability to inhibit TGase [Case, A., et al. (2005) Anal. Biochem. 338, 237-244]. In this paper, we report the kinetics of interaction of human TGase with one of the inhibitors that we identified, LDN-27219. We found that this compound is a reversible, slow-binding inhibitor that appears not to bind at the enzyme's active site but rather at the enzyme's GTP site, or a site that regulates binding of GTP. Interestingly, the potency and kinetics of inhibition are dependent on substrate structure and suggest a novel mechanism of inhibition that involves differential binding of LDN-27219 to multiple conformational states of this enzyme.     

High-resolution solution structure of Bacillus subtilis IIAglc

The high-resolution solution structure of the phosphocarrier protein IIA^glc from Bacillus subtilis is determined using 3D and 4D heteronuclear NMR methods. B. subtilis IIA^glc contains 162 amino acid residues and is one of the larger proteins for which high-resolution solution structure has been determined by NMR methods. The structures have been calculated from a total of 2,232 conformational constraints. Comparison with the X-ray crystal structure indicates that the overall fold is the same in solution and in crystalline environments, although some local structural differences are observed. These occur largely in turns and loops, and mostly correspond to regions with high-temperature factors in the crystal structure. The N-terminus of IIA^glc is disordered in solution. The active site is located in a concave region of the protein surface. The histidine, which accepts the phosphoryl group (His 83), interacts with a neighboring histidine (His 68) and is surrounded by hydrophobic residues. Proteins 31:258–270, 1998. © 1998 Wiley-Liss, Inc.     

Impacts of Argentine ants on avian nesting success

Biological invasions can have severe and widespread impacts on ecological communities. A few species of ants have become particularly damaging invaders but quantitative data of their impacts on many taxa is still lacking. We provide experimental evidence using artificial nests baited with quail eggs that the invasive Argentine ant (Linepithema humile) can be a significant avian nest predator – Argentine ants recruited to more nests and in higher abundance than the native ant species they displace. However, at a site invaded by Argentine ants, we monitored over 400 nests of a ground-nesting species, the Dark-eyed Junco (Junco hyemalis), and found that less than 2% of nests failed as a result of Argentine ant predation/infestation. A review of the literature also suggests that Argentine ants may not be a serious threat to bird nests relative to other predators or parasites. However, invasive ants with the capability of overwhelming prey though stinging (specifically the red-imported fire ant, Solenopsis invicta), may have a higher impact on avian nesting success. Received 14 January 2005; revised 28 April 2005; accepted 12 May 2005.     

Type IV Secretion-Dependent Activation of Host MAP Kinases Induces an Increased Proinflammatory Cytokine Response to Legionella pneumophila

The immune system must discriminate between pathogenic and nonpathogenic microbes in order to initiate an appropriate response. Toll-like receptors (TLRs) detect microbial components common to both pathogenic and nonpathogenic bacteria, whereas Nod-like receptors (NLRs) sense microbial components introduced into the host cytosol by the specialized secretion systems or pore-forming toxins of bacterial pathogens. The host signaling pathways that respond to bacterial secretion systems remain poorly understood. Infection with the pathogen Legionella pneumophila, which utilizes a type IV secretion system (T4SS), induced an increased proinflammatory cytokine response compared to avirulent bacteria in which the T4SS was inactivated. This enhanced response involved NF-κB activation by TLR signaling as well as Nod1 and Nod2 detection of type IV secretion. Furthermore, a TLR- and RIP2-independent pathway leading to p38 and SAPK/JNK MAPK activation was found to play an equally important role in the host response to virulent L. pneumophila. Activation of this MAPK pathway was T4SS-dependent and coordinated with TLR signaling to mount a robust proinflammatory cytokine response to virulent L. pneumophila. These findings define a previously uncharacterized host response to bacterial type IV secretion that activates MAPK signaling and demonstrate that coincident detection of multiple bacterial components enables immune discrimination between virulent and avirulent bacteria.     

Structure–activity relationship study of EphB3 receptor tyrosine kinase inhibitors

A structure–activity relationship study for a 2-chloroanilide derivative of pyrazolo[1,5-a]pyridine revealed that increased EphB3 kinase inhibitory activity could be accomplished by retaining the 2-chloroanilide and introducing a phenyl or small electron donating substituents to the 5-position of the pyrazolo[1,5-a]pyridine. In addition, replacement of the pyrazolo[1,5-a]pyridine with imidazo[1,2-a]pyridine was well tolerated and resulted in enhanced mouse liver microsome stability. The structure–activity relationship for EphB3 inhibition of both heterocyclic series was similar. Kinase inhibitory activity was also demonstrated for representative analogs in cell culture. An analog (32, LDN-211904) was also profiled for inhibitory activity against a panel of 288 kinases and found to be quite selective for tyrosine kinases. Overall, these studies provide useful molecular probes for examining the in vitro, cellular and potentially in vivo kinase-dependent function of EphB3 receptor.     

Frequency and recency of infection and their relationship with disgust and contamination sensitivity

Both disgust and contamination sensitivity likely evolved to protect us from infectious disease. Paradoxically, disgust may be reduced by frequent exposure to disgust-inducing cues — cues most likely to occur in disease-rich environments. In this study, we examined whether more frequent or recent illness might act to reverse this process. To test this, we surveyed 616 adults, obtaining illness frequency and recency data, disgust and contamination sensitivity, and a variety of control measures. Heightened contamination sensitivity was associated with more frequent infectious illness, but not with recency of infection. We also found that participants who had heightened contamination sensitivity and who were also more disgust sensitive had significantly fewer recent infections. These findings suggest that frequent illness may up-regulate contamination sensitivity potentially counteracting the effects of exposure on disgust. More importantly, these data provide the first direct evidence of a protective effect of contamination and disgust, against infectious disease.     

Sex differences in obesity rates in poor countries: Evidence from South Africa

Globally, men and women face markedly different risks of obesity. In all but of handful of (primarily Western European) countries, obesity is much more prevalent among women than men. We examine several potential explanations for this phenomenon. We analyze differences between men and women in reports and effects of potential underlying causes of obesity—childhood and adult poverty, depression, and attitudes about obesity. We evaluate the evidence for each explanation using data collected in an urban African township in the Cape Town metropolitan area. Three factors explain the greater obesity rates we find among women. Women who were nutritionally deprived as children are significantly more likely to be obese as adults, while men who were deprived as children face no greater risk. In addition, women of higher adult socioeconomic status are significantly more likely to be obese, which is not true for men. These two factors - childhood circumstances and adult SES - can fully explain the difference in obesity rates between men and women that we find in our sample. More speculatively, in South Africa, women's perceptions of an ‘ideal’ female body are larger than men's perceptions of the ‘ideal’ male body, and individuals with larger ‘ideal’ body images are significantly more likely to be obese.     

Generation of an OMgp allelic series in mice

The very limited ability to regenerate axons after injury in the mature mammalian central nervous system (CNS) has been partly attributed to the growth restrictive nature of CNS myelin. Oligodendrocyte myelin glycoprotein (OMgp) was identified as a major myelin‐derived inhibitor of axon growth. However, its role in axon regeneration in vivo is poorly understood. Here we describe the generation and molecular characterization of an OMgp allelic series. With a single gene targeting event and Cre/FLP mediated recombination, we generated an OMgp null allele with a LacZ reporter, one without a reporter gene, and an OMgp conditional allele. This allelic series will aid in the study of OMgp in adult CNS axon regeneration using mouse models of spinal cord injury. The conditional allele will overcome developmental compensation when employed with an inducible Cre, and allows for the study of temporal and tissue/cell type‐specific roles of OMgp in CNS injury‐induced axonal plasticity. genesis 47:751–756, 2009. © 2009 Wiley‐Liss, Inc.