The role of residues R97 and Y331 in modulating the pH optimum of an insect beta-glycosidase of family 1.

The activity of the digestive beta-glycosidase from Spodoptera frugiperda (Sfbetagly50, pH optimum 6.2) depends on E399 (pKa = 4.9; catalytic nucleophile) and E187 (pKa = 7.5; catalytic proton donor). Homology modelling of the Sfbetagly50 active site confirms that R97 and Y331 form hydrogen bonds with E399. Site-directed mutagenesis showed that the substitution of R97 by methionine or lysine increased the E399 pKa by 0.6 or 0.8 units, respectively, shifting the pH optima of these mutants to 6.5. The substitution of Y331 by phenylalanine increased the pKa of E399 and E187 by 0.7 and 1.6 units, respectively, and displaced the pH optimum to 7.0. From the observed deltapKa it was calculated that R97 and Y331 contribute 3.4 and 4.0 kJ.mol(-1), respectively, to stabilization of the charged E399, thus enabling it to be the catalytic nucleophile. The substitution of E187 by D decreased the pKa of residue 187 by 0.5 units and shifted the pH optimum to 5.8, suggesting that an electrostatic repulsion between the deprotonated E399 and E187 may increase the pKa of E187, which then becomes the catalytic proton donor. In short the data showed that a network of noncovalent interactions among R97, Y331, E399 and E187 controls the Sfbetagly50 pH optimum. As those residues are conserved among the family 1 beta-glycosidases, it is proposed here that similar interactions modulate the pH optimum of all family 1 beta-glycosidases.     

Floodplain and upland populations of Amazonian Himatanthus sucuuba: Effects of flooding on germination, seedling growth and mortality

Himatanthus sucuuba is a tree species that occurs in Central Amazonian white water floodplains and in non-flooded uplands. The objective of this paper was to verify whether flooding causes differences in germination percentages, seedling mortality as well as alterations of leaf number and biomass increments, related to the habitat of origin of the seeds (várzea floodplain versus terra firme upland, in the following V and TF, respectively). To test this effect, seeds from populations of both habitats were collected, and germination percentages were calculated under different experimental conditions. One hundred seeds collected in three sites of the floodplain and one hundred seeds from three sites of the upland were placed in trays for germination in a greenhouse in the Amazon Research Institute (INPA) in Manaus, Brazil. Seedlings with a height of 7 cm were then subjected to a period of 120 days of flooding in tanks with three treatments: control, waterlogging (root system and the base of the stem), and complete submersion. Ground water from the INPA was used in the tanks and was changed at weekly intervals. Germination percentages were high in most treatments. Seedling mortality was significantly higher in seedlings originating from upland, reaching 100% in submerged seedlings. On the other hand, várzea-originated seedlings showed no mortality under waterlogging or submergence.

Height growth of waterlogged seedlings from floodplains exceeded that of the control. Submerged seedlings did not grow, independently of the origin, and those originating from uplands died after about 100 days. Number of leaves and leaf area were significantly lower under partial submersion than in the control, independently of the origin of the seeds. Under total submersion abscission of all leaves was verified 30 days after inundation for seedlings of both origins. In the V population, all seedlings survived the waterlogged period, whereas in the TF population, 30% of the seedlings died when subjected to waterlogging. With complete submergence, in the V population 30% of the seedlings had died after 120 days, in the TF population after 90 days mortality was 100%. The results show that germination, seedling growth and seedling survival in H. sucuuba differ among populations of floodplain and upland sites: V plants performed better than TF plants under waterlogging and under submersion. The results strongly indicate that ecotypic differentiation exists between these two populations of H. sucuuba individuals in the two habitat types, which may be enforced by a strong selective pressure due to the regular occurring flood pulse.     

Effects of progressive and maximal exercise on plasma levels of adhesion molecules in athletes with sickle cell trait with or without alpha-thalassemia.

The aim of the study was to examine the effects of exercise on soluble vascular cell adhesion molecule-1 (sVCAM-1) and intercellular adhesion molecule-1 (sICAM-1) in sickle cell trait (SCT) athletes with or without alpha-thalassemia. Six athletes with SCT, seven athletes with both SCT and alpha-thalassemia (SCTAT), and seven control athletes (Cont) performed an incremental and maximal test on cycloergometer. Levels of sICAM-1 and sVCAM-1 were assessed at rest, immediately after the end of exercise, and 1, 2, and 24 h after exercise. Although Cont and SCTAT groups exhibited similar basal plasma levels of inflammatory and adhesion molecules, the SCT group had higher sVCAM-1 basal concentrations. Incremental exercise resulted in a significant increase of sVCAM-1 in all subjects, which remained elevated only in the SCT group during the recovery period. In conclusion, as sVCAM-1 increased with exercise and during the recovery period, our findings support the concept that SCT athletes might be at risk for microcirculatory disturbances and adhesive phenomena developing at rest and several hours after exercise. alpha-Thalassemia might be considered protective among exercising SCT subjects.     

Dynamics of noninvasively estimated microvascular O2 extraction during ramp exercise.

Utilization of near-infrared spectroscopy (NIRS) in clinical exercise testing to detect microvascular abnormalities requires characterization of the responses in healthy individuals and theoretical foundation for data interpretation. We examined the profile of the deoxygenated hemoglobin signal from NIRS {deoxygenated hemoglobin + myoglobin [deoxy-(Hb+Mb)] approximately O(2) extraction} during ramp exercise to test the hypothesis that the increase in estimated O(2) extraction would be close to hyperbolic, reflecting a linear relationship between muscle blood flow (Q(m)) and muscle oxygen uptake (Vo(2)(m)) with a positive Q(m) intercept. Fifteen subjects (age 24 +/- 5 yr) performed incremental ramp exercise to fatigue (15-35 W/min). The deoxy-(Hb+Mb) response, measured by NIRS, was fitted by a hyperbolic function [f(x) = ax/(b + x), where a is the asymptotic value and b is the x value that yields 50% of the total amplitude] and sigmoidal function {f(x) = f(0) + A/[1 + e(-(-c+dx))], where f(0) is baseline, A is total amplitude, and c is a constant dependent on d, the slope of the sigmoid}, and the goodness of fit was determined by F test. Only one subject demonstrated a hyperbolic increase in deoxy-(Hb+Mb) (a = 170%, b = 193 W), whereas 14 subjects displayed a sigmoidal increase in deoxy-(Hb+Mb) (f(0) = -7 +/- 7%, A = 118 +/- 16%, c = 3.25 +/- 1.14, and d = 0.03 +/- 0.01). Computer simulations revealed that sigmoidal increases in deoxy-(Hb+Mb) reflect a nonlinear relationship between microvascular Q(m) and Vo(2)(m) during incremental ramp exercise. The mechanistic implications of our findings are that, in most healthy subjects, Q(m) increased at a faster rate than Vo(2)(m) early in the exercise test and slowed progressively as maximal work rate was approached.     

Mechanical Complications of Myocardial Infarction—Radiologic Aspects

Left ventricular aneurysm, interventricular septal defect and acute mitral valve incompetence due to papillary muscle damage are three mechanical complications which cause intractable heart failure following myocardial infarction. In each case surgical intervention can result in dramatic improvement of congestive heart failure.A hemodynamically significant left ventricular aneurysm enlarges the cardiac silhouette and frequently causes a localized protrusion as seen radiographically. Cardiac fluoroscopy will disclose an abnormal pulsation of the left ventricular border. The left ventricular angiogram establishes the diagnosis, reveals the extent of the aneurysm and may disclose a filling defect in the aneurysmal sac due to the presence of mural thrombus. Coronary arteriography shows occlusion of a major vessel, most commonly the anterior descending branch of the left coronary artery.Ischemic perforation of the interventricular septum and acute mitral incompetence due to severe papillary muscle damage both cause severe heart failure shortly after myocardial infarction. A similar pansystolic murmur accompanies both conditions, and differentiation between the two is rarely possible on the basis of the electrocardiogram or x-ray film of the chest. Ventricular cardiac catheterization and left ventricular angiocardiography are required for a correct diagnosis.     

Burkholderia cepacia lipase: A versatile catalyst in synthesis reactions

The lipase from Burkholderia cepacia, formerly known as Pseudomonas cepacia lipase, is a commercial enzyme in both soluble and immobilized forms widely recognized for its thermal resistance and tolerance to a large number of solvents and short‐chain alcohols. The main applications of this lipase are in transesterification reactions and in the synthesis of drugs (because of the properties mentioned above). This review intends to show the features of this enzyme and some of the most relevant aspects of its use in different synthesis reactions. Also, different immobilization techniques together with the effect of various compounds on lipase activity are presented. This lipase shows important advantages over other lipases, especially in reaction media including solvents or reactions involving short‐chain alcohols.     

Floral resources and risk of exposure to pesticides for Melipona quadrifasciata anthidioides Lepeletier 1836 in a Cerrado of São Paulo (Brazil)

Honey and bee bread samples from storage pots of Melipona quadrifasciata anthidioides were collected monthly from April 2015 to May 2016 in the Mogi Guaçu Biological Reserve (22º 10? S, 47º 11? W). The flora in the site is characteristic of the Atlantic Forest with preserved areas of savanna-like vegetation surrounded by commercial forests, orchards and various crops of exotic and native plants. Samples were analysed with the use of melissopalynological methodology and 46 pollen types from 38 genera and 30 families were identified in 25 honey samples. Fabaceae, Asteraceae, Myrtaceae, Sapindaceae showed the greatest pollen richness in honey. Predominant nectariferous pollen types were Anadenanthera, Cordia, Eucalyptus, Mimosa scabrella, Schefflera, Sida, Serjania and Vernonia. Twenty-eight types of pollen from 21 genera and 19 families were identified in 22 bee bread samples. Fabaceae, Asteraceae and Myrtaceae showed the highest pollen richness. Anadenanthera, Cecropia, Eucalyptus, Melastomataceae, Mimosa scabrella, Mimosa verrucosa and Myrcia were the most frequent polliniferous pollen types. Principal component analysis (PCA) demonstrated that honey and pollen samples formed two main groups of similarity, mainly due to Eucalyptus’ nectar and pollen of Melastomataceae, respectively. Melipona quadrifasciata anthidioides collected nectar and pollen from the preserved areas as well as in the secondary and ‘ruderal’ vegetation and in cultivated forests/fields, suggesting their importance as pollinators both of native flora and exotic species. The use of trophic resources of plants grown with pesticides is a concern for the conservation of these species of bee and should be better studied.