Pheromone binding by polymorphic mouse major urinary proteins

Mouse major urinary proteins (MUPs) have been proposed to play a role in regulating the release and capture of pheromones. Here, we report affinity measurements of five recombinant urinary MUP isoforms (MUPs-I, II, VII, VIII, and IX) and one recombinant nasal isoform (MUP-IV) for each of three pheromonal ligands, (+/-)-2-sec-butyl-4,5-dihydrothiazole (SBT), 6-hydroxy-6-methyl-3-heptanone (HMH), and (+/-)dehydro-exo-brevicomin (DHB). Dissociation constants for all MUP-pheromone pairs were determined by isothermal titration calorimetry, and data for SBT were corroborated by measurements of intrinsic protein fluorescence. We also report the isolation of MUP-IV protein from mouse nasal extracts, in which MUP-IV mRNA has been observed previously. The affinity of each MUP isoform for SBT (K(d) approximately 0.04 to 0.9 micro M) is higher than that for DHB (K(d) approximately 26 to 58 micro M), which in turn is higher than that for HMH (K(d) approximately 50 to 200 micro M). Isoforms I, II, VIII, and IX show very similar affinities for each of the ligands. MUP-VII has approximately twofold higher affinity for SBT but approximately twofold lower affinity for the other pheromones, whereas MUP-IV has approximately 23-fold higher affinity for SBT and approximately fourfold lower affinity for the other pheromones. The variations in ligand affinities of the MUP isoforms are consistent with structural differences in the binding cavities of the isoforms. The data indicate that the concentrations of available pheromones in urine may be influenced by changes in the expression levels of urinary MUPs or the excretion levels of other MUP ligands. The variation in pheromone affinities of the urinary MUP isoforms provides only limited support for the proposal that MUP heterogeneity plays a role in regulating profiles of available pheromones. However, the binding data support the proposed role of nasal MUPs in sequestering pheromones and possibly transporting them to their receptors.     

Isolation and characterization of highly liganded protein from brewer's barley grain

Two basic proteins, HLP-1 and HLP-2, were isolated from brewer's barley grain (Hordeum vulgare L.) and characterized as glycoproteins with molecular masses of 16 and 13 kDa and pI values of 7.4 and 8.8, respectively. They could bind sugars, metal ions, and both hydrophobic and hydrophylic molecules of low molecular mass. These characteristics may be related to their potential plant-protecting role.     

A-hydroxyphosphonate oligonucleotides: a promising DNA type?

The synthesis of monomers (S)-1, (R)-1 and 2 derived from (5'S)-, (5'R)-2'-deoxythymidine-5'-C-phosphonic acids and 2',5'-dideoxythymidine-5'-C-phosphonic acids was elaborated. The protection of the 5'-hydroxyl by the methoxycarbonyl group was a key step of the synthesis. Prepared monomers were used for the solid-phase assembly of several types oligothymidylate 15-mers (S)-3, (S)-4, (S)-5, (R)-4 and (R)-5 containing the chiral 3'-O-P-CH(OH)-5' internucleotide linkage. Their hybridization properties with dA15 and rA15 were studied as well as their resistance against nuclease cleavage.     

Structural,dynamic and mechanical properties of POPC at low cholesterol concentration studied in pressure/temperature space

We have studied the structural, dynamic and mechanical properties of 1-palmitoyl-2-oleoyl- sn-glycero-3-phosphatidylcholine (POPC)/cholesterol binary mixtures by small-angle X-ray scattering. Our investigations were concentrated on the biologically most relevant pressure-temperature-cholesterol regime, i.e. the liquid crystalline phase and its phase boundary to the lamellar gel phase within a cholesterol concentration up to 25 mol%. From the dependence of the transition pressure we derived a value of 19 kJ/mol for the transition enthalpy Delta H(m) of POPC in excess water. With increasing cholesterol concentration, Delta H(m) drops to about 7 kJ/mol at 20 mol% cholesterol. Time-resolved pressure-scan (p-scan) and temperature-jump (T-jump) experiments reveal that at low cholesterol content (<5-8 mol%) the fluidity and also the bilayer compressibility increase remarkably. In contrast, at concentrations between 5 and 25 mol% cholesterol the bilayer becomes again more rigid and the lipid bilayer spacing increases about 2 A. Theses changes are attributed to the onset of phase separation between liquid disordered and liquid ordered phases. The fluid-fluid miscibility gap for this mono-unsaturated lecithin species is strongly enlarged compared with saturated lecithins.     

Three-dimensional spine kinematics during multidirectional,target-directed trunk movement in sitting

The current study provides a quantitative assessment of three-dimensional spine motion during target-directed trunk movements in sitting. Subjects sat on an elevated surface, without foot support, and targets were placed in five directions, at three subject-specific distances (based on trunk height). Subjects were asked to lean toward the target, touch it with their head, and return to upright sitting. A retro-reflective motion analysis system was used to measure spine motion, using three kinematic trunk models (1, 3 and 7 segments). Significant differences were noted in the total trunk motion measured between the models, as well as between target distances and directions. In the most segmented model, inter-segmental trunk motion was also found to differ between trunk levels, with complex interaction effects involving target distance and direction. These findings suggest that inter-segmental spine motion is complex, task dependent, and often unevenly distributed between spine levels, with motion patterns differing between subjects, even in the absence of pathology. Use of a multi-segmental model provides the most interpretable findings, allowing for differentiation of individual motion patterns of the spine. Such an approach may be beneficial to the understanding of movement-related spine pathologies.     

Optimal combination of minimum degrees of freedom to be actuated in the lower limbs to facilitate arm-free paraplegic standing

Arm-free paraplegic standing via functional electrical stimulation (FES) has drawn much attention in the biomechanical field as it might allow a paraplegic to stand and simultaneously use both arms to perform daily activities. However, current FES systems for standing require that the individual actively regulates balance using one or both arms, thus limiting the practical use of these systems. The purpose of the present study was to show that actuating only six out of 12 degrees of freedom (12-DOFs) in the lower limbs to allow paraplegics to stand freely is theoretically feasible with respect to multibody stability and physiological torque limitations of the lower limb DOF. Specifically, the goal was to determine the optimal combination of the minimum DOF that can be realistically actuated using FES while ensuring stability and able-bodied kinematics during perturbed arm-free standing. The human body was represented by a three-dimensional dynamics model with 12-DOFs in the lower limbs. Nakamura's method (Nakamura, Y., and Ghodoussi, U., 1989, "Dynamics Computation of Closed-Link Robot Mechanisms With Nonredundant and Redundant Actuators," IEEE Trans. Rob. Autom., 5(3), pp. 294-302) was applied to estimate the joint torques of the system using experimental motion data from four healthy subjects. The torques were estimated by applying our previous finding that only 6 (6-DOFs) out of 12-DOFs in the lower limbs need to be actuated to facilitate stable standing. Furthermore, it was shown that six cases of 6-DOFs exist, which facilitate stable standing. In order to characterize each of these cases in terms of the torque generation patterns and to identify a potential optimal 6-DOF combination, the joint torques during perturbations in eight different directions were estimated for all six cases of 6-DOFs. The results suggest that the actuation of both ankle flexionextension, both knee flexionextension, one hip flexionextension, and one hip abductionadduction DOF will result in the minimum torque requirements to regulate balance during perturbed standing. To facilitate unsupported FES-assisted standing, it is sufficient to actuate only 6-DOFs. An optimal combination of 6-DOFs exists, for which this system can generate able-bodied kinematics while requiring lower limb joint torques that are producible using contemporary FES technology. These findings suggest that FES-assisted arm-free standing of paraplegics is theoretically feasible, even when limited by the fact that muscles actuating specific DOFs are often denervated or difficult to access.     

14-3-3 expression in denervated hippocampus after entorhinal cortex lesion assessed by culture-derived isotope tags in quantitative proteomics

Activation of astrocytes accompanies many brain pathologies. Reactive astrocytes have a beneficial role in acute neurotrauma but later on might inhibit regeneration. 2D-gel electrophoresis and mass spectrometry were applied to study the proteome difference in denervated hippocampus in wildtype mice and mice lacking intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin (GFAP-/-Vim-/-) that show attenuated reactive gliosis and enhanced posttraumatic regeneration. Proteomic data and immunohistochemical analyses showed upregulation of the adapter protein 14-3-3 four days postlesion and suggested that 14-3-3 upregulation after injury is triggered by reactive gliosis. Culture-derived isotope tags (CDIT) and mass spectrometry demonstrated that 14-3-3 epsilon was the major isoform upregulated in denervated hippocampus and that its upregulation was attenuated in GFAP-/-Vim-/- mice and thus most likely connected to reactive gliosis.     

Chemical composition and antimicrobial activity of the essential oil of endemic Dalmatian black pine (Pinus nigra ssp. dalmatica)

The chemical composition and the antimicrobial activity of the essential oil isolated from the needles of endemic Dalmatian black pine (Pinus nigra ssp. dalmatica) from Croatia were investigated. The chemical composition of the essential oil was determined by GC and GC/MS analyses, and the main compounds identified were α-pinene, β-pinene, germacrene D, and β-caryophyllene. Disc-diffusion and broth-microdilution assays were used for the in vitro antimicrobial screening. The Dalmatian black pine essential oil exhibited a great potential of antibacterial activity against Gram-positive bacteria (MIC=0.03-0.50% (v/v)) and a less pronounced activity against Gram-negative bacteria (MIC=0.12-3.2% (v/v)). The volatile compounds also inhibited the growth of all fungi tested, including yeast.