A Universal Scaffold for Synthesis of the Fe(CN)2(CO) Moiety of [NiFe] Hydrogenase

Hydrogen-cycling [NiFe] hydrogenases harbor a dinuclear catalytic center composed of nickel and iron ions, which are coordinated by four cysteine residues. Three unusual diatomic ligands in the form of two cyanides (CN⁻) and one carbon monoxide (CO) are bound to the iron and apparently account for the complexity of the cofactor assembly process, which involves the function of at least six auxiliary proteins, designated HypA, -B, -C, -D, -E, and -F. It has been demonstrated previously that the HypC, -D, -E, and -F proteins participate in cyanide synthesis and transfer. Here, we show by infrared spectroscopic analysis that the purified HypCD complexes from Ralstonia eutropha and Escherichia coli carry in addition to both cyanides the CO ligand. We present experimental evidence that in vivo the attachment of the CN⁻ ligands is a prerequisite for subsequent CO binding. With the aid of genetic engineering and subsequent mutant analysis, the functional role of conserved cysteine residues in HypD from R. eutropha was investigated. Our results demonstrate that the HypCD complex serves as a scaffold for the assembly of the Fe(CN)₂(CO) entity of [NiFe] hydrogenase.     

Morphological respiratory diffusion capacity of the lungs of ball pythons (Python regius)

This study aims at a functional and morphological characterization of the lung of a boid snake. In particular, we were interested to see if the python's lungs are designed with excess capacity as compared to resting and working oxygen demands. Therefore, the morphological respiratory diffusion capacity of ball pythons (Python regius) was examined following a stereological, hierarchically nested approach. The volume of the respiratory exchange tissue was determined using computed tomography. Tissue compartments were quantified using stereological methods on light microscopic images. The tissue diffusion barrier for oxygen transport was characterized and measured using transmission electron micrographs. We found a significant negative correlation between body mass and the volume of respiratory tissue; the lungs of larger snakes had relatively less respiratory tissue. Therefore, mass-specific respiratory tissue was calculated to exclude effects of body mass. The volume of the lung that contains parenchyma was 11.9±5.0mm(3)g(-1). The volume fraction, i.e., the actual pulmonary exchange tissue per lung parenchyma, was 63.22±7.3%; the total respiratory surface was, on average, 0.214±0.129m(2); it was significantly negatively correlated to body mass, with larger snakes having proportionally smaller respiratory surfaces. For the air-blood barrier, a harmonic mean of 0.78±0.05μm was found, with the epithelial layer representing the thickest part of the barrier. Based on these findings, a median diffusion capacity of the tissue barrier ( [Formula: see text] ) of 0.69±0.38ml O(2)min(-1)mmHg(-1) was calculated. Based on published values for blood oxygen concentration, a total oxygen uptake capacity of 61.16mlO(2)min(-1)kg(-1) can be assumed. This value exceeds the maximum demand for oxygen in ball pythons by a factor of 12. We conclude that healthy individuals of P. regius possess a considerable spare capacity for tissue oxygen exchange.     

Genetic and biochemical characterization of the cell wall hydrolase activity of the major secreted protein of Lactobacillus rhamnosus GG

Lactobacillus rhamnosus GG (LGG) produces two major secreted proteins, designated here Msp1 (LGG_00324 or p75) and Msp2 (LGG_00031 or p40), which have been reported to promote the survival and growth of intestinal epithelial cells. Intriguingly, although each of these proteins shares homology with cell wall hydrolases, a physiological function that correlates with such an enzymatic activity remained to be substantiated in LGG. To investigate the bacterial function, we constructed knock-out mutants in the corresponding genes aiming to establish a genotype to phenotype relation. Microscopic examination of the msp1 mutant showed the presence of rather long and overly extended cell chains, which suggests that normal daughter cell separation is hampered. Subsequent observation of the LGG wild-type cells by immunofluorescence microscopy revealed that the Msp1 protein accumulates at the septum of exponential-phase cells. The cell wall hydrolyzing activity of the Msp1 protein was confirmed by zymogram analysis. Subsequent analysis by RP-HPLC and mass spectrometry of the digestion products of LGG peptidoglycan (PG) by Msp1 indicated that the Msp1 protein has D-glutamyl-L-lysyl endopeptidase activity. Immunofluorescence microscopy and the failure to construct a knock-out mutant suggest an indispensable role for Msp2 in priming septum formation in LGG.     

Lipoteichoic acid is an important microbe-associated molecular pattern of Lactobacillus rhamnosus GG

Background

Receptors with a single transmembrane (TM) domain are essential for the signal transduction across the cell membrane. NMR spectroscopy is a powerful tool to study structure of the single TM domain. The expression and purification of a TM domain in Escherichia coli (E.coli) is challenging due to its small molecular weight. Although ketosteroid isomerase (KSI) is a commonly used affinity tag for expression and purification of short peptides, KSI tag needs to be removed with the toxic reagent cyanogen bromide (CNBr).

Result

The purification of the TM domain of p75 neurotrophin receptor using a KSI tag with the introduction of a thrombin cleavage site is described herein. The recombinant fusion protein was refolded into micelles and was cleaved with thrombin. Studies showed that purified protein could be used for structural study using NMR spectroscopy.

Conclusions

These results provide another strategy for obtaining a single TM domain for structural studies without using toxic chemical digestion or acid to remove the fusion tag. The purified TM domain of p75 neurotrophin receptor will be useful for structural studies.     

The Head and Neck Anatomy of Sea Turtles (Cryptodira: Chelonioidea) and Skull Shape in Testudines

Background

Sea turtles (Chelonoidea) are a charismatic group of marine reptiles that occupy a range of important ecological roles. However, the diversity and evolution of their feeding anatomy remain incompletely known.

Methodology/Principal Findings

Using computed tomography and classical comparative anatomy we describe the cranial anatomy in two sea turtles, the loggerhead (Caretta caretta) and Kemp’s ridley (Lepidochelys kempii), for a better understanding of sea turtle functional anatomy and morphological variation. In both taxa the temporal region of the skull is enclosed by bone and the jaw joint structure and muscle arrangement indicate that palinal jaw movement is possible. The tongue is relatively small, and the hyoid apparatus is not as conspicuous as in some freshwater aquatic turtles. We find several similarities between the muscles of C. caretta and L. kempii, but comparison with other turtles suggests only one of these characters may be derived: connection of the m. adductor mandibulae internus into the Pars intramandibularis via the Zwischensehne. The large fleshy origin of the m. adductor mandibulae externus Pars superficialis from the jugal seems to be a characteristic feature of sea turtles.

Conclusions/Significance

In C. caretta and L. kempii the ability to suction feed does not seem to be as well developed as that found in some freshwater aquatic turtles. Instead both have skulls suited to forceful biting. This is consistent with the observation that both taxa tend to feed on relatively slow moving but sometimes armoured prey. The broad fleshy origin of the m. adductor mandibulae externus Pars superficialis may be linked to thecheek region being almost fully enclosed in bone but the relationship is complex.     

Web search queries can predict stock market volumes

We live in a computerized and networked society where many of our actions leave a digital trace and affect other people's actions. This has lead to the emergence of a new data-driven research field: mathematical methods of computer science, statistical physics and sociometry provide insights on a wide range of disciplines ranging from social science to human mobility. A recent important discovery is that search engine traffic (i.e., the number of requests submitted by users to search engines on the www) can be used to track and, in some cases, to anticipate the dynamics of social phenomena. Successful examples include unemployment levels, car and home sales, and epidemics spreading. Few recent works applied this approach to stock prices and market sentiment. However, it remains unclear if trends in financial markets can be anticipated by the collective wisdom of on-line users on the web. Here we show that daily trading volumes of stocks traded in NASDAQ-100 are correlated with daily volumes of queries related to the same stocks. In particular, query volumes anticipate in many cases peaks of trading by one day or more. Our analysis is carried out on a unique dataset of queries, submitted to an important web search engine, which enable us to investigate also the user behavior. We show that the query volume dynamics emerges from the collective but seemingly uncoordinated activity of many users. These findings contribute to the debate on the identification of early warnings of financial systemic risk, based on the activity of users of the www.     

Temporal bone arrangements in turtles: an overview

The temporal region of turtles is characterized by significant anatomical diversity. Turtles show a pure anapsid morphotype that exhibits various different marginal reductions known as emarginations. As a result of this diversity, turtles can be taken as a model by which to understand the processes that may have resulted in the highly debated anatomy of the amniote temporal region in general. In this review on almost forgotten literature, I summarize ten potential factors that may act on the skull to shape the temporal region of turtles. These are: (1) phylogenetic constraints, (2) skull weights, (3) type of food, (4) skull dimensions, (5) muscle bulging, (6) ear anatomy and jaw muscle bending mechanisms, (7) extent and nature of muscle attachment sites, (8) internal forces within the jaw adductor chamber, (9) environmental pressure, and (10) neck bending mechanisms. Particular focus is laid on the interrelationship of the jaw musculature and the dermatocranial armour, which were assumed to influence each other to a certain degree. In the literature, cranial dimensions were assumed to influence temporal bone formation within major tetrapod groups. Among these, turtles seem to represent a kind of intermixture, a phenomenon that may be reflected in their specific anatomy. The references presented should be understood as product of the scientific environment in which they developed and the older literature does not always insist current empirical demands. However, the intuitive and creative ideas and the comprehensive anatomical considerations of these authors may inspire future studies in several fields related to this topic.