Unfolding of preproteins upon import into mitochondria.

Unfolding of preproteins and translocation across the mitochondrial membranes requires their interaction with mt-Hsp70 and Tim44 at the inner face of the inner membrane and ATP as an energy source. We measured the temperature dependence of the rates of unfolding and import into the matrix of two folded passenger domains, the tightly folded heme-binding domain (HBD) of cytochrome b2 and the loosely folded mouse dihydrofolate reductase (DHFR). Despite the stability of the HBD, its rates of thermal breathing were fast and the preprotein was imported rapidly at all temperatures. In contrast, rates of unfolding and import of DHFR were strongly temperature dependent and import was significantly slower than unfolding. In addition, import rates of DHFR were strongly dependent on the length of the presequence. We propose that the mitochondrial import motor does not exert a constant pulling force. Rather, mt-Hsp70 appears to release a translocating polypeptide chain such that the precursor can then slide back and refold on the surface of the mitochondria. Refolding competes with translocation, and passengers may undergo several rounds of unfolding and refolding prior to their import.     

Where Failure Is Not an Option –Personalized Medicine in Astronauts

Drug safety and efficacy are highly variable among patients. Most patients will experience the desired drug effect, but some may suffer from adverse drug reactions or gain no benefit. Pharmacogenetic testing serves as a pre-treatment diagnostic option in situations where failure or adverse events should be avoided at all costs. One such situation is human space flight. On the international space station (ISS), a list of drugs is available to cover typical emergency settings, as well as the long-term treatment of common conditions for the use in self-medicating common ailments developing over a definite period. Here, we scrutinized the list of the 78 drugs permanently available at the ISS (year 2014) to determine the extent to which their metabolism may be affected by genetic polymorphisms, potentially requiring genotype-specific dosing or choice of an alternative drug. The purpose of this analysis was to estimate the potential benefit of pharmacogenetic diagnostics in astronauts to prevent therapy failure or side effects.     

SAR of N-phenyl piperidine based oral integrin α5β1 antagonists

Recently, a new class of selective integrin α5β1inhibitors consisting of a heterocyclic based scaffold was published. Herein the SAR and pharmacokinetic profiles of N-phenyl piperidine derivatives are described.     

Relevance of glycine and cysteine residues as well as N- and C-terminals for the activity of protein histidine phosphatase

There is increasing evidence that reversible phosphorylation of histidine residues regulates numerous important cellular processes. The first protein histidine phosphatase (PHP) from vertebrates was discovered just recently. Here, we report on amino acids and domains essential for activity of PHP. Point mutations of conserved residues and deletions of the N- and C-termini of PHP were analyzed using [³²P-his]ATP-citrate lyase as a substrate. Individual or joint replacement of all cysteine residues by alanine did not affect PHP activity. Deletion of 9 N-terminal amino acids resulted in inactive PHP. Furthermore, only 4 C-terminal residues could be deleted without losing PHP activity. Single or multiple mutations of the glycine-rich domain (Gly⁷⁵, Gly⁷⁷) of a putative nucleotide binding site of PHP (GxGxxG/S) caused inactivation of PHP. Wildtype PHP could be labeled with [α-³²P]ATP. Such radiolabeling was not detectable for catalytically inactive PHP-G75A and PHP-G77A. These data suggest further studies on the interaction between PHP and ATP.     

Effects of Polyphosphate Additives on Campylobacter Survival in Processed Chicken Exudates

Campylobacter spp. are responsible for a large number of the bacterial food poisoning cases worldwide. Despite being sensitive to oxygen and nutritionally fastidious, Campylobacter spp. are able to survive in food processing environments and reach consumers in sufficient numbers to cause disease. To investigate Campylobacter persistence on processed chicken, exudates from chickens produced for consumer sale were collected and sterilized. Two types of exudates from chicken products were collected: enhanced, where a marinade was added to the chickens during processing, and nonenhanced, where no additives were added during processing. Exudates from enhanced chicken products examined in this study contained a mixture of polyphosphates. Exudate samples were inoculated with Campylobacter jejuni or Campylobacter coli strains and incubated under a range of environmental conditions, and viable bacteria present in the resultant cultures were enumerated. When incubated at 42°C in a microaerobic environment, exudates from enhanced chicken products resulted in increased survival of C. jejuni and C. coli compared with that in nonenhanced exudates in the range of <1 to >4 log CFU/ml. Under more relevant food storage conditions (4°C and normal atmosphere), the exudates from enhanced chicken products also demonstrated improved Campylobacter survival compared with that in nonenhanced exudates. Polyphosphates present in the enhanced exudates were determined to be largely responsible for the improved survival observed when the two types of exudates were compared. Therefore, polyphosphates used to enhance chicken quality aid in sustaining the numbers of Campylobacter bacteria, increasing the opportunity for disease via cross-contamination or improperly cooked poultry.Campylobacter species are the major causative agent of food-borne gastrointestinal bacterial infections in the developed world (6, 11, 21). Poultry products are a major source for the introduction of Campylobacter into the food supply (15, 16). Improperly cooked poultry and cross-contamination of other foods by raw poultry are common methods for transmission of Campylobacter to humans (5). However, Campylobacter spp. are nutritionally fastidious organisms that are sensitive to the oxygen levels present in a normal environment (O₂ = 20.9%) (21). Therefore, Campylobacter appears an unlikely candidate to persist within poultry processing and storage environments at levels sufficient to cause human disease. This conundrum directly leads to a question: what then are the elements that contribute to the ability of Campylobacter to survive through poultry processing and cold storage?To investigate this question, a food-relevant environment consisting of chicken weepage or exudate can be used to perform survival experiments on Campylobacter species. Strains of Campylobacter jejuni and Campylobacter coli were used for the survival studies since these two species are responsible for the vast majority of human cases of campylobacteriosis (20, 28). Chicken exudate is the fluid that seeps out from processed poultry carcasses and is often found to be contaminated with considerable numbers of Campylobacter bacteria. It is comprised of water, blood, fats, and other materials added to the poultry during processing. Sterilized poultry exudates make for a convenient experimental material that is also relevant to the conditions which Campylobacter will experience as a contaminant of processed poultry (2, 3). Two different types of chicken exudates were collected from commercial producers, one from chickens processed without additives (nonenhanced) and the other from chickens that were treated with a commercial marinade to increase the quality and appeal of the meat at market (enhanced). The commercial poultry marinades contain a significant amount of polyphosphate additives. Polyphosphates comprise a group of food additives that are utilized within poultry processing to enhance the moisture absorbance, color, and flavor and to reduce product shrinkage of poultry (24, 29-32). Polyphosphates have also been shown to have an antimicrobial effect on several different bacterial species (8, 10, 12). The goal of the research was to determine if polyphosphates contribute to the ability of Campylobacter to survive and persist through the supply chain, thus directly increasing the opportunity for Campylobacter-mediated food poisoning of consumers.     

New species of the Hygrobates salamandrarum-group (Acari, Hydrachnidia, Hygrobatidae) from Southeast Asia

Two new water mite species of the genus Hygrobates Koch, 1837 (Acari, Hydrachnidia, Hygrobatidae) were found to live parasitic on newts of the genus Paramesotriton Chang, 1935 (Amphibia, Caudata, Salamandridae) from Vietnam and Laos: Hygrobates forcipifer sp. nov. and H. ancistrophorus sp. nov. The H. salamandrarum-group is defined, that now includes three species from Southeast Asia. Males and females of both new species are described, as well as larvae and deutonymphs of the Vietnamese species. These data provide the first record of males, nymphs and larvae of the species-group. The systematic position of the group, as well as the parasite-host association and the lifecycle of the species are discussed. Furthermore, the character states of the striking mouthparts, particularly modified as an adaptation for penetrating the amphibian skin, the genital skeleton and the larval morphology are examined.     

Analysis of methylarginine metabolism in the cardiovascular system identifies the lung as a major source of ADMA

Protein arginine methylation is catalyzed by a family of enzymes called protein arginine methyltransferases (PRMTs). Three forms of methylarginine have been identified in eukaryotes: monomethylarginine (l-NMMA), asymmetric dimethylarginine (ADMA), and symmetric dimethylarginine (SDMA), all characterized by methylation of one or both guanidine nitrogen atoms of arginine. l-NMMA and ADMA, but not SDMA, are competitive inhibitors of all nitric oxide synthase isoforms. SDMA is eliminated almost entirely by renal excretion, whereas l-NMMA and ADMA are further metabolized by dimethylarginine dimethylaminohydrolase (DDAH). To explore the interplay between methylarginine synthesis and degradation in vivo, we determined PRMT expression and DDAH activity in mouse lung, heart, liver, and kidney homogenates. In addition, we employed HPLC-based quantification of protein-incorporated and free methylarginine, combined with immunoblotting for the assessment of tissue-specific patterns of arginine methylation. The salient findings of the present investigation can be summarized as follows: 1) pulmonary expression of type I PRMTs was correlated with enhanced protein arginine methylation; 2) pulmonary ADMA degradation was undertaken by DDAH1; 3) bronchoalveolar lavage fluid and serum exhibited almost identical ADMA/SDMA ratios, and 4) kidney and liver provide complementary routes for clearance and metabolic conversion of circulating ADMA. Together, these observations suggest that methylarginine metabolism by the pulmonary system significantly contributes to circulating ADMA and SDMA levels.     

Bicarbonate and active site zinc modulate the self-peroxidation of bovine copper-zinc superoxide dismutase

Peroxidation reactions of copper-zinc superoxide dismutase (CuZn-SOD1) or its zinc-depleted form (CuE-SOD1) that likely also involve a component of bicarbonate buffer have been implicated in the pathophysiology of the neurodegenerative diseases amyotrophic lateral sclerosis (ALS), Alzheimer's Disease and Parkinson's Disease. Neither removal of the zinc ion nor adding bicarbonate had large effects on the self-peroxidation reaction of bovine SOD1, but the combination of zinc-deficiency and added bicarbonate caused major changes to the spin trapped SOD1-centred free radical. Removal of the active site zinc ion greatly decreased the formation of an unassigned SOD1-centred free radical in the reaction with the inorganic peroxide peroxynitrite. The results suggest that under cellular conditions ( approximately 5 mM bicarbonate) zinc-deficient SOD1 peroxidation could play a pathogenic role in neurodegenerative diseases.     

Combining ontogenetic and evolutionary scales of morphological disparity: a study of early Jurassic ammonites

Two major research themes in Evolutionary Developmental Biology and in Paleobiology, respectively, have each become central for the analysis and interpretation of morphological changes in evolution: the study of ontogeny/phylogeny connections, mainly within the widespread and controversial framework of heterochrony; and the study of morphological disparity, the morphological signal of biodiversity, describing secular changes in morphospace occupation during the history of any given clade. Although enriching in their respective fields, these two themes have remained rather isolated to date, despite the potential value of integrating them as some recent studies begin to suggest. Here, we explore the recent notion of developmental morphospace-morphospace carrying ontogenetic information-as a potential tool for bridging the gap between disparity dynamics and developmental dynamics. We elaborate this approach with a case study of Early Jurassic ammonite family Hildoceratidae (Mollusca, Cephalopoda). Morphometric analyses of the shell shape of 20 species spanning the morphological spectrum of the family are used to quantify and contrast juvenile and adult disparity levels. Adult disparity is significantly greater than juvenile disparity at the family level; yet, some subclades also display different patterns. In addition, comparisons of ontogenetic trajectories underline the prevalence of heterochrony-based evolutionary modifications within subfamilies (via ontogenetic scaling); they also point to the probable existence of pervasive developmental constraints structuring inhomogeneous morphospace occupation.