Iodination of proteins by IPy2BF4, a new tool in protein chemistry

Iodination is a very useful method for protein characterization and labeling. However, derivatization chemistries used in most conventional iodination procedures may cause substantial alterations in protein structure and function. The IPy(2)BF(4) reagent [bis(pyridine)iodonium (I) tetrafluoroborate] has been shown to be an effective iodinating reagent for peptides. Herein we report the first application of IPy(2)BF(4) in protein iodination in an aqueous medium using three representative substrates: insulin, lysozyme, and the enzyme 1,3-1,4-beta-d-4-glucanohydrolase. Our results show that IPy(2)BF(4) has clear advantages over existing methods in that the reaction is quantitative, fast, and selective for the most accessible Tyr residues of a protein, and it preserves the functional integrity of the protein when moderate Tyr labeling levels are pursued.     

To the heart of myofibril assembly

One of the most fascinating examples of cytoskeletal assembly is the myofibril, the contractile structure of striated (i.e. skeletal and cardiac) muscle. Myofibrils are composed of repeating contractile units known as sarcomeres, perhaps the most highly ordered macromolecular structures in eukaryotic cells. When skeletal and cardiac muscle cells differentiate, thousands of structural and regulatory molecules assemble into the semicrystalline sarcomeric contractile units. As a consequence of this precise assembly, many different classes of proteins function together to convert the molecular interactions of actin and myosin efficiently into the macroscopic movements of contractile activity.     

Cooperative vigilance: the guanaco's (Lama guanicoe) key antipredator mechanism

The concept of sociality has been associated with the effectiveness of antipredator mechanisms, like cooperative vigilance and the dilution effect. Lama guanicoe (guanaco) is a social native herbivore in South America and a social species. The objectives of this study were to evaluate the antipredator responses of different-sized groups of guanacos in areas with varying predation risks and to determine antipredator mechanisms in guanacos. For this, we measured different antipredator responses to a potential predator (human subjects). Detection of predator and flight distances from predator both increased with a greater number of guanacos per group and with greater distances among guanacos within the social group. Both buffer distance and flight time decreased with a greater number of guanacos per group, but increased with greater distances among guanacos inside the social group. Solitary adult males moved shorter distance and mixed groups moved greater distances. Flight distances were greater in areas with tall and dense vegetation than in areas with low vegetation. Buffer distance and flight time were shorter in undulating land than on flat lands, and groups were usually observed on hill slopes. Our results suggest that the benefit of social grouping in guanacos is the increased probability of avoiding predator with cooperative vigilance and not with the dilution effect. This means that a predator could be detected earlier when approaching a guanaco group than when approaching a solitary individuals and could thus be avoided.     

Growth hormone response induced by a respiratory muscle endurance training in healthy subjects

To date, the large majority of studies evaluating growth hormone (GH) response to acute physical exercise has been performed involving gross muscle groups. To the best of our knowledge, none has evaluated the effects of a respiratory muscle endurance training (RMET) on hormonal secretions, particularly on GH release, though some respiratory devices have been widely used in athletes to train respiratory muscles and to improve cardiopulmonary function and physical performance. 8 healthy men underwent an incremental progressive RMET protocol of 11 daily sessions, obtained through the use of a specifically designed respiratory device (Spiro Tiger?). The 12th session of RMET (15 min duration: 1 min at a respiration rate of 28 acts/min, 5 min at 32 acts/min, 5 min at 34 acts/min, 4 min at 36 acts/min) was associated with blood samplings for determination of GH, cortisol, ghrelin, glucose, and lactate (LA) levels. GH and cortisol responses significantly increased after a 15-minute RMET session, which, in contrast, inhibited ghrelin secretion. There was a minimal, though significant, increase in LA levels with a significant elevation in glycemia. A 15-minute RMET session, administered after a 11-days incremental progressive RMET protocol, was capable of stimulating GH and cortisol release and suppressing ghrelin secretion. Optimization of incremental progressive RMET protocols would be important to maximize the positive chronic effects of this intervention on somatotropic function and muscle performance.     

Shared resistance to aging and ALS in neuromuscular junctions of specific muscles

Normal aging and neurodegenerative diseases both lead to structural and functional alterations in synapses. Comparison of synapses that are generally similar but respond differently to insults could provide the basis for discovering mechanisms that underlie susceptibility or resistance to damage. Here, we analyzed skeletal neuromuscular junctions (NMJs) in 16 mouse muscles to seek such differences. We find that muscles respond in one of three ways to aging. In some, including most limb and trunk muscles, age-related alterations to NMJs are progressive and extensive during the second postnatal year. NMJs in other muscles, such as extraocular muscles, are strikingly resistant to change. A third set of muscles, including several muscles of facial expression and the external anal sphinter, succumb to aging but not until the third postnatal year. We asked whether susceptible and resistant muscles differed in rostrocaudal or proximodistal position, source of innervation, motor unit size, or fiber type composition. Of these factors, muscle innervation by brainstem motor neurons correlated best with resistance to age-related decline. Finally, we compared synaptic alterations in normally aging muscles to those in a mouse model of amyotrophic lateral sclerosis (ALS). Patterns of resistance and susceptibility were strikingly correlated in the two conditions. Moreover, damage to NMJs in aged muscles correlated with altered expression and distribution of CRMP4a and TDP-43, which are both altered in motor neurons affected by ALS. Together, these results reveal novel structural, regional and molecular parallels between aging and ALS.     

Immobilization of Escherichia coli containing ω-transaminase activity in LentiKats®

Whole Escherichia coli cells overexpressing ω‐transaminase (ω‐TA) and immobilized cells entrapped in LentiKats® were used as biocatalysts in the asymmetric synthesis of the aromatic chiral amines 1‐phenylethylamine (PEA) and 3‐amino‐1‐phenylbutane (APB). Whole cells were permeabilized with different concentrations of cetrimonium bromide (CTAB) and ethanol; the best results were obtained with CTAB 0.1% which resulted in an increase in reaction rate by 40% compared to the whole cells. The synthesis of PEA was carried out using isopropyl amine (IPA) and L ‐alanine (Ala) as amino donors. Using whole cell biocatalysis, the reaction with IPA was one order of magnitude faster than with Ala. No reaction was detected when permeabilized E. coli cells containing ω‐TA were employed using Ala as the amino donor. Additionally, the synthesis of APB from 4‐phenyl‐2‐butanone and IPA was studied. Whole and permeabilized cells containing ω‐TA and their immobilized LentiKats® counterparts showed similar initial reactions rates and yields in the reaction systems, indicating 100% of immobilization efficiency (observed activity/activity immobilized) and absence of diffusional limitations (due to the immobilization). Immobilization of whole and permeabilized cells containing ω‐TA in LentiKats® allowed improved stability as the biocatalyst was shown to be efficiently reused for five reaction cycles, retaining around 80% of original activity. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Time of Progression to Osteopenia/Osteoporosis in Chronically HIV-Infected Patients: Screening DXA Scan

Background

Algorithms for bone mineral density (BMD) management in HIV-infected patients are lacking. Our objective was to assess how often a dual-energy x-ray absorptiometry (DXA) scan should be performed by assessing time of progression to osteopenia/osteoporosis.

Methods

All DXA scans performed between 2000 and 2009 from HIV-infected patients with at least two DXA were included. Time to an event (osteopenia and osteoporosis) was assessed using the Kaplan–Meier method. Strata (tertiles) were defined using baseline minimum T scores. Differences between strata in time to an event were compared with the log-rank test.

Results

Of 391 patients (1,639 DXAs), 49.6% had osteopenia and 21.7% osteoporosis at their first DXA scan. Of the 112 (28.6%) with normal BMD, 35.7% progressed to osteopenia; median progression time was 6.7 years. These patients were stratified: “low-risk" (baseline minimum T score >−0.2 SD), “middle-risk" (between −0.2 and −0.6 SD), and “high-risk" (from −0.6 to −1 SD); median progression time to osteopenia was 8.7, >7.2, and 1.7 years, respectively (p<0.0001). Of patients with osteopenia, 23.7% progressed to osteoporosis; median progression time was >8.5 years. Progression time was >8.2 years in “low-risk" tertile (T score between −1.1 and −1.6 SD), >8.5 years in “middle-risk" (between −1.6 and −2), and 3.2 years in “high-risk" (from −2 to −2.4) (p<0.0001).

Conclusions

Our results may help to define the BMD testing interval. The lowest T score tertiles would suggest recommending a subsequent DXA in 1–2 years; in the highest tertiles, ≥6 years. Early intervention in patients with bone demineralization could reduce fracture–related morbidity/mortality.     

Effect of Cattle on Salmonella Carriage,Diversity and Antimicrobial Resistance in Free-Ranging Wild Boar (Sus scrofa) in Northeastern Spain

Salmonella is distributed worldwide and is a pathogen of economic and public health importance. As a multi-host pathogen with a long environmental persistence, it is a suitable model for the study of wildlife-livestock interactions. In this work, we aim to explore the spill-over of Salmonella between free-ranging wild boar and livestock in a protected natural area in NE Spain and the presence of antimicrobial resistance. Salmonella prevalence, serotypes and diversity were compared between wild boars, sympatric cattle and wild boars from cattle-free areas. The effect of age, sex, cattle presence and cattle herd size on Salmonella probability of infection in wild boars was explored by means of Generalized Linear Models and a model selection based on the Akaike’s Information Criterion. Prevalence was higher in wild boars co-habiting with cattle (35.67%, CI 95% 28.19–43.70) than in wild boar from cattle-free areas (17.54%, CI 95% 8.74–29.91). Probability of a wild boar being a Salmonella carrier increased with cattle herd size but decreased with the host age. Serotypes Meleagridis, Anatum and Othmarschen were isolated concurrently from cattle and sympatric wild boars. Apart from serotypes shared with cattle, wild boars appear to have their own serotypes, which are also found in wild boars from cattle-free areas (Enteritidis, Mikawasima, 4:b:- and 35:r:z35). Serotype richness (diversity) was higher in wild boars co-habiting with cattle, but evenness was not altered by the introduction of serotypes from cattle. The finding of a S. Mbandaka strain resistant to sulfamethoxazole, streptomycin and chloramphenicol and a S. Enteritidis strain resistant to ciprofloxacin and nalidixic acid in wild boars is cause for public health concern.