Thiol-activated serine proteinases from nymphal hemolymph of the African migratory locust,Locusta migratoria migratorioides

Two unique serine proteinase isoenzymes (LmHP-1 and LmHP-2) were isolated from the hemolymph of African migratory locust (Locusta migratoria migratorioides) nymphs. Both have a molecular mass of about 23 kDa and are activated by thiol-reducing agents. PMSF abolishes enzymes activity only after thiol activation, while the cysteine proteinase inhibitors E-64, iodoacetamide, and heavy metals fail to inhibit the thiol-activated enzymes. The N-terminal sequence was determined for the more-abundant LmHP-2 isoenzyme. It exhibits partial homology to that of other insect serine proteinases and similar substrate specificity and inhibition by the synthetic and protein trypsin inhibitors pABA, TLCK, BBI, and STI. The locust trypsins LmHP-1 and LmHP-2 constitute a new category of serine proteases wherein the active site of the enzyme is exposed by thiol activation without cleavage of peptide bonds.     

Biochemical, Physiological, and Pathological Aspects of the Peripheral Benzodiazepine Receptor

The PBR is a mitochondrial protein composed of at least two subunits, an approximately 30-kDa subunit that contains the site for BZs and an approximately 18-kDa subunit that binds isoquinoline carboxamide derivatives. Porphyrins and diazepam binding inhibitor are putative endogenous ligands for these receptors, which are under neural and hormonal control. Alterations in the density of PBR seem to be a sensitive indicator of stress: up-regulation after acute stress and down-regulation induced by repeated stress. PBR-specific ligands are involved in the control of cell proliferation and differentiation, and their binding is increased in some cancer tumors. Numerous studies in various endocrine organs have revealed that PBR are located in specific regions or tissues in the organs. Furthermore, PBR densities in various organs subject to hormonal control are regulated by organotropic hormones. At least in some cases, BZ ligands do not exert a specific effect in an organ, but rather modulate the well-documented effects of that particular hormone. To the best of our knowledge, BZ ligand action in peripheral tissues is dependent on recognition of PBR, which may suggest a receptor-mediated action.     

The action pattern and physiological role of Tenebrio larval amylase

The larval midgut amylase of Tenebrio molitor L. may be classified as a typical -amylase on the basis of its action pattern on amylopectin, glycogen, and β-amylase limit dextrin. When the development is on the preferred diet of wheat-bran, the amylatic activity of the larvae is impaired by an -amylase inhibitor present in wheat. The inhibitor has been found to be stable to digestion by the larvae. Wheat β-amylase plays the major role in the digestion of starch by larvae. The implications of these relations are discussed.     

31P and 2H relaxation studies of helix VII and the cytoplasmic helix of the human cannabinoid receptors utilizing solid-state NMR techniques

Cannabinoid receptors are G-protein-coupled receptors comprised of seven transmembrane helices. We hypothesized that the extended helix of the receptor interacts differently with POPC bilayers due to the differing distribution of charged amino acid residues. To test this, hCB1(T377-E416) and hCB2(K278-H316) peptides were studied with 31P and 2H solid-state NMR spectroscopy by incorporating them into 1-palmitoyl-2-oleoyl-sn-glycerophosphocholine bilayers. Lipid affinities of the 40- and 39-residue peptides were analyzed on the basis of 31P and 2H spectral line shapes, order parameters, and T1 relaxation measurements of the POPC bilayers. Lipid headgroup perturbations were noticed in the 31P NMR spectra in the lipid/peptide mixtures when compared with the pure lipids. 2H order parameters were calculated from the quadrupolar splitting of the de-Paked 2H NMR spectra. At the top of the acyl chain, pure lipids had an average S(CD) approximately = 0.20, whereas S(CD) approximately = 0.16 and S(CD) approximately = 0.18 were found in the presence of hCB1(T377-E416) and hCB2(K278-H316), respectively. S(CD) values decreased in the central part of the acyl chains when compared to the pure POPC lipids, indicating a change in the dynamic properties of the lipid membrane in the presence of the cannabinoid peptides. R(1Z) vs S2(CD) plots exhibited a linear dependency with and without the peptides, with an increase in slope upon addition of the peptides to the POPC, indicating that the dynamics of the lipid bilayer is dominated by fast axially symmetric motion. This study provides insights into the interaction of cannabinoid peptides with the membrane bilayer by investigating the headgroup and acyl chain dynamics.     

The structural context of disease-causing mutations in gap junctions

Gap junctions form intercellular channels that mediate metabolic and electrical signaling between neighboring cells in a tissue. Lack of an atomic resolution structure of the gap junction has made it difficult to identify interactions that stabilize its transmembrane domain. Using a recently computed model of this domain, which specifies the locations of each amino acid, we postulated the existence of several interactions and tested them experimentally. We introduced mutations within the transmembrane domain of the gap junction-forming protein connexin that were previously implicated in genetic diseases and that apparently destabilized the gap junction, as evidenced here by the absence of the protein from the sites of cell-cell apposition. The model structure helped identify positions on adjacent helices where second-site mutations restored membrane localization, revealing possible interactions between residue pairs. We thus identified two putative salt bridges and one pair involved in packing interactions in which one disease-causing mutation suppressed the effects of another. These results seem to reveal some of the physical forces that underlie the structural stability of the gap junction transmembrane domain and suggest that abrogation of such interactions bring about some of the effects of disease-causing mutations.     

Respiratory‐induced vasoconstriction measured by light transmission and by laser Doppler signal

Changes in finger tissue blood volume (TBV) measured by light transmission and in laser Doppler flow (LDF) were obtained during long breathing (of 12 s period) and associated with the respiratory phases, inspiration and expiration. For fifteen out of sixteen subjects TBV and LDF started to decrease 0–2 s after the start of expiration and increased during inspiration but the start of increase occurred before the start of inspiration, showing that the respiratory‐induced changes in TBV and LDF are mainly associated with the expiration. Decrease of TBV and LDF after expiration was also found during the inspiratory gasps (© 2013 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Protein synthesis and secretion by the locust fat body after ovariectomy

The synthesis and secretion of proteins, including vitellogenin, by the locust fat body were investigated in vivo and in vitro at various times after ovariectomy. The rate of overall protein synthesis and secretion by the fat body in vivo is significantly less in ovariectomized animals than in the controls two to three weeks after the operation, the difference in the rates of secretion of vitellin antibody-precipitable protein being even more pronounced and detectable earlier. The significantly greater amount of newly synthesized vitellogenin retained in the fat body of ovariectomized animals is insufficient to account for this difference. The secretion of newly synthesized protein by the fat bodies of ovariectomized locusts in vitro is significantly less than that of control fat bodies, the difference being particularly marked in the case of vitellogenin. The polysome populations of fat bodies of ovariectomized and control females are quantitatively similar and the amounts of total protein and vitellin antibody-precipitable protein synthesized by these polysomes in a cell-free system do not differ significantly.     

Fine‐root exploitation strategies differ in tropical old growth and logged‐over forests in Ghana

Understanding the changes in root exploitation strategies during post‐logging recovery is important for predicting forest productivity and carbon dynamics in tropical forests. We sampled fine (diameter < 2 mm) roots using the soil core method to quantify fine‐root biomass and architectural and morphological traits to determine root exploitation strategies in an old growth forest and in a 54‐yr‐old logged‐over forest influenced by similar parent material and climate. Seven root traits were considered: four associated with resource exploitation potential or an ‘extensive’ strategy (fine‐root biomass, length, surface area, and volume), and three traits which reflect exploitation efficiency or an ‘intensive’ strategy (specific root area, specific root length, and root tissue density). We found that total fine‐root biomass, length, surface area, volume, and fine‐root tissue density were higher in the logged‐over forest, whereas the old growth forest had higher total specific root length and specific root surface area than the logged‐over forest. The results suggest different root exploitation strategies between the forests. Plants in the old growth forest invest root biomass more efficiently to maximize soil volume explored, whereas plants in the logged‐over forest increase the spatial distribution of roots resulting in the expansion of the rhizosphere.