Structure of a complex of human lactoferrin N-lobe with pneumococcal surface protein a provides insight into microbial defense mechanism

Human lactoferrin, a component of the innate immune system, kills a wide variety of microorganisms including the Gram positive bacteria Streptococcus pneumoniae. Pneumococcal surface protein A (PspA) efficiently inhibits this bactericidal action. The crystal structure of a complex of the lactoferrin-binding domain of PspA with the N-lobe of human lactoferrin reveals direct and specific interactions between the negatively charged surface of PspA helices and the highly cationic lactoferricin moiety of lactoferrin. Binding of PspA blocks surface accessibility of this bactericidal peptide preventing it from penetrating the bacterial membrane. Results of site-directed mutagenesis, in vitro protein binding assays and isothermal titration calorimetry measurements corroborate that the specific electrostatic interactions observed in the crystal structure represent major associations between PspA and lactoferrin. The structure provides a snapshot of the protective mechanism utilized by pathogens against the host's first line of defense. PspA represents a major virulence factor and a promising vaccine candidate. Insights from the structure of the complex have implications for designing therapeutic strategies for treatment and prevention of pneumococcal diseases that remain a major public health problem worldwide.     

A new method to determine the electrical transfer function of the human thorax

Traditional analyses have assumed that cardiac electrical activity is reflected on the surface ECG without distortion as the signal passes through the body tissues. This study aims to explore the frequency dependence of thoracic attenuation of surface-recorded intracardiac electrical activity. Twenty patients (14 men, 55 +/- 15 yr of age) undergoing electrophysiological study were enrolled. Rectangular unipolar stimuli were applied from a catheter positioned in the right ventricular apical area and another in the posteroseptal area without contact with the myocardium. An orthogonal Frank-lead surface ECG and a unipolar intracardiac electrogram near the pacing site were recorded. Frequency domain characteristics of the signal-averaged pacing impulses were analyzed. Linear regression analysis showed significant frequency-dependent attenuation in the magnitude transfer functions (R(2) = 0.84-0.89, P < 0.0001) and good linear fit for the phase transfer characteristics (R(2) = 0.98-1.0, P < 0.0001). Age, physical dimension, and respiratory characteristics had significant effects on the magnitude and phase characteristics of the transfer functions. Application of models of the low- and high-slope transfer functions to signal-averaged ECGs from 33 subjects showed differences in the attenuation of P and T waves relative to the QRS.     

Structural basis for ubiquitin-mediated dimerization and activation of the ubiquitin protein ligase Cbl-b

Cbl proteins are E3 ubiquitin ligases that are negative regulators of many receptor tyrosine kinases. Cbl-b and c-Cbl contain a ubiquitin-associated (UBA) domain, which is present in a variety of proteins involved in ubiquitin-mediated processes. Despite high sequence identity, Cbl UBA domains display remarkably different ubiquitin-binding properties. Here, we report the crystal structure of the UBA domain of Cbl-b in complex with ubiquitin at 1.9 A resolution. The structure reveals an atypical mechanism of ubiquitin recognition by the first helix of the UBA. Helices 2 and 3 of the UBA domain form a second binding surface, which mediates UBA dimerization in the crystal and in solution. Site-directed mutagenesis demonstrates that Cbl-b dimerization is regulated by ubiquitin binding and required for tyrosine phosphorylation of Cbl-b and ubiquitination of Cbl-b substrates. These studies demonstrate a role for ubiquitin in regulating biological activity by promoting protein dimerization.     

Extracellular amino acid levels in the human liver during transplantation: a microdialysis study from donor to recipient

Using microdialysis, we have monitored extracellular levels of amino acids and related amines in the human liver at three stages of the transplantation procedure: donor retrieval, back table preparation and during 48 h post-implantation. By comparing the ratio of mean levels at the donor and back table stages, with the ratio between early (2-6 h) and late (43-48 h) post-reperfusion, these amines were classified into one of three groups. In one group, back table levels were markedly higher than during the donor stage, with levels declining over time post-reperfusion. A second group had much lower levels in the back table than during the donor phase, and post-reperfusion levels were either stable or increased over time. Concentrations of amino acids in the final group remained relatively constant at all stages. This study illustrates the value of microdialysis in providing organ-specific metabolic data that may indicate specific mechanisms of poor graft function.     

Evolution of an intermediate C4 photosynthesis in the non-foliar tissues of the Poaceae

Photosynthesis Research - Carbon concentrating mechanisms (CCMs) in plants are abaptive features that have evolved to sustain plant growth in unfavorable environments, especially at low atmospheric...     

Conformational regulation of the EGFR kinase core by the juxtamembrane and C-terminal tail: a molecular dynamics study

The catalytic domain of epidermal growth factor receptor (EGFR) is activated by dimerization, which requires allosteric coupling between distal dimerization and catalytic sites. Although crystal structures of EGFR kinases, solved in various conformational states, have provided important insights into EGFR activation by dimerization, the atomic details of how dimerization signals are dynamically coupled to catalytic regions of the kinase core are not fully understood. In this study, we have performed unrestrained and targeted molecular dynamics simulations on the active and inactive states of EGFR, followed by principal component analysis on the simulated trajectories, to identify correlated motions in the EGFR kinase domain upon dimerization. Our analysis reveals that the conformational changes associated with the catalytic functions of the kinase core are highly correlated with motions in the juxtamembrane (JM) and C-terminal tail, two flexible structural elements that play an active role in EGFR kinase activation and dimerization. In particular, the opening and closing of the ATP binding lobe relative to the substrate binding lobe is highly correlated with motions in the JM and C-terminal tail, suggesting that ATP and substrate binding can be coordinated with dimerization through conformational changes in the JM and C-terminal tail. Our study pinpoints key residues involved in this conformational coupling, and provides new insights into the role of the JM and C-terminal tail segments in EGFR kinase functions.     

Plasma free amino acid kinetics in rainbow trout (<Emphasis Type="Italic">Oncorhynchus mykiss</Emphasis>) using a bolus injection of <Superscript>15</Superscript>N-labeled amino acids

To gain insight into the metabolic design of the amino acid carrier systems in fish, we injected a bolus of ¹⁵N amino acids into the dorsal aorta in mature rainbow trout (Oncorhynchus mykiss). The plasma kinetic parameters including concentration, pool size, rate of disappearance (R _d), half-life and turnover rate were determined for 15 amino acids. When corrected for metabolic rate, the R _d values obtained for trout for most amino acids were largely comparable to human values, with the exception of glutamine (which was lower) and threonine (which was higher). R _d values ranged from 0.9 μmol 100 g⁻¹ h⁻¹ (lysine) to 22.1 μmol 100 g⁻¹ h⁻¹ (threonine) with most values falling between 2 and 6 μmol 100 g⁻¹ h⁻¹. There was a significant correlation between R _d and the molar proportion of amino acids in rainbow trout whole body protein hydrolysate. Other kinetic parameters did not correlate significantly with whole body amino acid composition. This indicates that an important design feature of the plasma-free amino acids system involves proportional delivery of amino acids to tissues for protein synthesis.     

Evidences of lateral gene transfer between archaea and pathogenic bacteria

Acquisition of new genetic material through horizontal gene transfer has been shown to be an important feature in the evolution of many pathogenic bacteria. Changes in the genetic repertoire, occurring through gene acquisition and deletion, are the major events underlying the emergence and evolution of bacterial pathogens. However, horizontal gene transfer across the domains i.e. archaea and bacteria is not so common. In this context, we explore events of horizontal gene transfer between archaea and bacteria. In order to determine whether the acquisition of archaeal genes by lateral gene transfer is an important feature in the evolutionary history of the pathogenic bacteria, we have developed a scheme of stepwise eliminations that identifies archaeal-like genes in various bacterial genomes. We report the presence of 9 genes of archaeal origin in the genomes of various bacteria, a subset of which is also unique to the pathogenic members and are not found in respective non-pathogenic counterparts. We believe that these genes, having been retained in the respective genomes through selective advantage, have key functions in the organism’s biology and may play a role in pathogenesis.