Oxygen equilibrium ofPriapulus hemerythrin

Summary The hemerythrin-containing coelomic fluid ofPriapulus caudatus shows a relatively low O₂ affinity (half-saturation O₂ tension P₅₀=8 mm at 10 °C) and a low O₂ capacity (near 1 vol.%). O₂ affinity is independent of pH but shows a large temperature sensitivity. A major role as a continuous O₂ transporter seems to be excluded.Acknowledgments. A major part of this work was carried out at the Kristineberg Marine Laboratory, Fiskebäckskil (Sweden) and in the Zoophysiology Department, Aarhus Universitet (Denmark).     

Presynaptic effects of spider toxins: Increase of high affinity uptake in the arthropod peripheral glutamatergic system

In contrast to the reported effects of polyamines on the high affinity neurotransmitter uptake, two polyamine-like spider toxins significantly increase the high affinity uptake of glutamate as demonstrated with high resolution autoradiography. The effects of both spider toxins were compared to those of a polyamine toxin from the waspPhilanthus triangulum, which is known to inhibit the high affinity glutamate uptake.     

Preparation of a specific ligand for the purification of arylsulfatases by affinity chromatography

Summary The reduction ofp-nitrocatechol sulfate, the artificial substrate of arylsulfatases followed by a condensation of a succinic anhydride, gives a ligand for the purification of the enzyme by affinity chromatography.     

Involvement of a surface concanavalin A-binding glycoprotein in the adhesion ofTrichomonas vaginalis to substrates

Summary Treatment ofTrichomonas vaginalis with EDTA removes their ability to adhere to glass surfaces and changes their affinity to Concanavalin A (ConA) by a different distribution of their surface structures. Filtrates of the EDTA-treatedTrichomonas passed through affinity chromatography columns (ConA bound to Sepharose 4B0 separate into 2 fractions, one fraction was bound to the ConA-Sepharose beads, the other was not. The Con A-bound fraction appears to be a glycoprotein which restores in a specific way the ability of the EDTA-treated protozoa to adhere to glass. We want to thank Dr.P. M. Comoglio for the donation of the FITC-labelled immunoglobulin fraction of anti-ConA rabbit antiserum. This work was supported by a grant from the Italian National Research Council (C.N.R.)     

Binding of matrilysin-1 to human epithelial cells promotes its activity

Matrix metalloproteinase-7 (MMP-7, matrilysin- 1) modulates crucial biological events by processing many epithelial cell surface-associated effectors. We addressed MMP-7 interaction with human epithelial cells and its resulting activity. In human endometrium, a model of controlled tissue remodeling, proMMP-7 was diffusely immunolocalized inside epithelial cells, whereas MMP-7 delineated their entire plasma membrane. Endometrial explants preferentially retained active MMP-7, but not proMMP-7. Endometrial epithelial cells and carcinoma cells from various tissues bound active MMP-7. Endometrial carcinoma-derived Ishikawa cells showed high affinity (K_D of ~2.5 nM) and capacity (~260 000 sites per cell) for MMP-7. MMP-7 binding decreased by extracting membrane sterols or interfering with heparan sulfate proteoglycans, and was abrogated by tissue inhibitors of metalloproteinase-2 (TIMP-2) or synthetic MMP inhibitors. Bound MMP-7 not only remained fully active towards a macromolecular substrate but also became resistant to TIMP-2. We conclude that MMP-7-selective targeting to the plasma membrane of epithelial cells promotes its activity by conferring resistance to TIMP-2. A. Berton, C. Selvais: These authors contributed equally to this work. P. J. Courtoy, E. Marbaix, H. Emonard: These authors contributed equally to the supervision of this work. Received 20 September 2006; received after revision 30 November 2006; accepted 18 January 2007     

Blockade by zinc ions of K+-induced contraction and calcium in guinea pigTaenia coli

Preincubation with 0.3 mM Zn²⁺ markedly inhibited both the tonic response and Ca²⁺ binding at low affinity sites induced by K⁺ (60 mM), with smaller effects on the phasic response and the high affinity Ca²⁺ sites, inTaenia coli. However, when the muscle was kept in Zn²⁺-containing medium following the first stimulation with the K⁺, the phasic response and the high affinity Ca²⁺ sites were more severely inhibited during the second stimulation with K⁺. This probably indicates that Zn²⁺ reduced the tonic tension response to K⁺ mainly by inhibiting Ca²⁺ influx at the cell membranes ofTaenia coli. However, when Zn²⁺ is continuously present, Ca²⁺ is not supplied at the storage sites and is not available for the phasic response to a second stimulation with K⁺.     

A lectin in the pollen of marihuana,Cannabis sativa L.

Summary A lectin was found in the pollen ofCannabis sativa L. which reacted with papain treated erythrocytes. The lectin has an affinity for glucose containing receptor sites.     

Purification of an L-fucose binding lectin fromUlex europeus by affinity column chromatography

Summary An L-fucose binding lectin fromUlex europeus was purified by affinity column chromatography using an L-fucose-starch complex. The lectin thus purified had a mol.wt of 60,000, and consisted of 2 glycoprotein subunit with mol.wt 29,000 and 31,000, respectively.     

The toxicity of twoBacillus thuringiensis δ-endotoxins to gypsy moth larvae is inversely related to the affinity of binding sites on midgut brush border membranes for the toxins

Summary The-endotoxin fromBacillus thuringiensis subspecieskurstaki strain HD1-9 is almost 400 times more potent than the-endotoxin from strain HD-73 as a gypsy moth larvicide. The two-endotoxins compete for a high-affinity binding site on the brush border membrane of larval gypsy moth midguts. The affinity for the-endotoxin from strain HD-73 is much greater than the affinity for the-endotoxin from strain HD1-9.     

Morphology of a non-occluded virus isolated from citrus red mite,Panonychus citri

Summary The size and morphology of virus particles isolated from citrus red mite (Panonychus citri) are similar to those observed in thin sections. The similarity to the virus particles isolated fromOryctes rhinoceros suggests affinity to theBaculovirus group.Presently at USDA, ARS, Fruit and Vegetable Insect Laboratory, Vincennes, IN 47591.     

The lectin fromViscum album L. purification by biospecific affinity chromatography

Summary A lectin fromViscum album which specifically binds to D-galactose was isolated by affinity chromatography on O-lactosyl-, O-galactosyl-polycarylamide or hydrolized sepharose 4 B. Some serological and physicochemical properties of the agglutinin are reported.     

Relationship between the absolute configuration and the biological activity of juvenile hormone III

Summary The activity of the pure 10R (=natural) and 10S enantiomers of juvenile hormone III (JH III) was determined in 3 different bioassays, and the relative binding affinity of the 2 enantiomers to the haemolymph JH-binding protein of the cockroachNauphoeta cinerea was measured. In theGalleria wax test, a local morphogenetic assay, the 10R enantiomer was 5240 times more active than, the 10S enantiomer, 1Galleria unit corresponding to 0.42 pg of 10R-JH III as compared to 2.2 ng for 10S-JH III. In a systemic morphogenetic assay with the cockroachNauphoeta cinerea 380 times less 10R enantiomer was necessary in order to induce detectable juvenilisation (58 ng 10R and 22 g 10S) and in a systemic gonadotropic assay withNauphoeta cinerea 255 times less 10R was needed to induce vitellogenin synthesis in 50% of the insects (6.7 ng 10R and 1710 ng 10S). In the JH-binding protein assay 10R-JH III had an affinity for the JH-binding protein (lipophorin) which was approximately 46 times higher than that of 10S-JH III.     

Relationship between lectin-affinity granules in anuran embryos and formation of primordial germ cells

Summary Embryos of the anuran,Rana nigromaculata, contained granules with a specific affinity for Con A and GS-I. Larvae derived from embryos injected with these lectins had a noticeable reduction in both the number and size of primordial germ cells (PGCs). This observation suggests that the granules are somehow involved in the formation of PGCs.     

The kinetics of estrogen binding to rat alpha-fetoprotein

Fluid obtained from rat fetuses was utilized to characterize the affinity, number of binding sites, and the association and dissociation rate kinetics of the binding of estradiol and estrone to AFP. Statistical analysis demonstrated no differences when the values for the AFP-estradiol interaction were compared with those obtained for the ATP-estrone interaction. These data demonstrate that rat AFP specifically binds estradiol and estrone with a high capacity, high affinity, and similar binding kinetics.     

Respiratory function of blood and relative weight of certain organs in two species of shrew: Crocidura russula and Suncus etruscus (Soricidae mammals)]

A high oxygen capacity combined with a low oxygen affinity and a large Bohr effect achieve a high oxygen transport capacity in small shrews. The high metabolic rate (Suncus estruscus: 100 to 350 ml O2/kg. min) can be explained by the combination of the favorable blood parameters with a high relative heart weight (1,2% of body weight) and heart frequencies of 1,000 to 1,350 per minute.     

<Emphasis Type="Italic">Anopheles gambiae</Emphasis> odorant binding protein crystal complex with the synthetic repellent DEET: implications for structure-based design of novel mosquito repellents

Insect odorant binding proteins (OBPs) are the first components of the olfactory system to encounter and bind attractant and repellent odors emanating from various sources for presentation to olfactory receptors, which trigger relevant signal transduction cascades culminating in specific physiological and behavioral responses. For disease vectors, particularly hematophagous mosquitoes, repellents represent important defenses against parasitic diseases because they effect a reduction in the rate of contact between the vectors and humans. OBPs are targets for structure-based rational approaches for the discovery of new repellent or other olfaction inhibitory compounds with desirable features. Thus, a study was conducted to characterize the high resolution crystal structure of an OBP of Anopheles gambiae, the African malaria mosquito vector, in complex with N,N-diethyl-m-toluamide (DEET), one of the most effective repellents that has been in worldwide use for six decades. We found that DEET binds at the edge of a long hydrophobic tunnel by exploiting numerous non-polar interactions and one hydrogen bond, which is perceived to be critical for DEET’s recognition. Based on the experimentally determined affinity of AgamOBP1 for DEET (K _d of 31.3 μΜ) and our structural data, we modeled the interactions for this protein with 29 promising leads reported in the literature to have significant repellent activities, and carried out fluorescence binding studies with four highly ranked ligands. Our experimental results confirmed the modeling predictions indicating that structure-based modeling could facilitate the design of novel repellents with enhanced binding affinity and selectivity.     

Isolation and characterization of biologically active lectin from Korean mistletoe, Viscum album var. Coloratum

A mistletoe lectin was isolated from water extracts of Korean mistletoe, a subspecies of Viscum album, grown on Quercus mongolica using CM-Sepharose chromatography followed by an affinity chromatography on a concanavalin A-Sepharose column. The compound proved to be a mistletoe lectin II with D-galactose and N-acetyl-D-galactosamine specificity. Matrix-assisted laser desorption time-of-flight mass spectroscopy showed it to have an average molecular mass of 62.7 kDa and to consist of two subunits of 30.6 kDa and 32.5 kDa. It was a basic protein with isoelectric points of 9.4 and 9.6 by capillary isoelectric focusing and was cytotoxic to Molt4 cell. Received 17 November 1998; received after revision 3 March 1999; accepted 3 March 1999     

Immunochemical similarities among the hemolymph juvenile hormone binding proteins of moths

The hemolymph from various species of moths was analyzed for cross-reactivity with a panel of six monoclonal antibodies made against the hemolymph juvenile hormone binding protein ofManduca sexta. With the exception of one antibody, the immunoreactivity was limited to the sphingid family. One monoclonal antibody cross-reacted with a number of lepidopteran species; however, families such as Noctuidae and Pyralidae, known to have high affinity, low molecular weight juvenile hormone binding proteins, did not cross-react. Immunological cross-reactivity withManduca sexta juvenile hormone binding protein in several primitive moth families supports the current model of phylogenetic relationships in the order Lepidoptera.     

Purification of breast cancer resistance protein ABCG2 and role of arginine-482

Human ABCG2 was efficiently overexpressed in insect cell membranes, solubilized with 3-[(3-cholamidopropyl)dimethyl ammonio]-1-propanesulfonate, and purified through N-terminal hexahistidine tag. Its functionality was assessed by high vanadate-sensitive ATPase activity, and nucleotide-binding capacity. Interestingly, the R482T point mutation increased both maximal hydrolysis rate and affinity for MgATP, and lowered sensitivity to vanadate inhibition. Direct nucleotide binding, as monitored by quenching of intrinsic fluorescence, indicated a mutation-related preference for ATP over ADP. The R482T mutation only produced a limited change, if any, on the binding of drug substrates, indicating that methotrexate, on the one hand, and rhodamine 123 or doxorubicin, on the other hand, bound similarly to wild-type and mutant transporters whether or not they were subject to cellular transport. In addition, the characteristic inhibitors GF120918 and 6-prenylchrysin, which alter mitoxantrone efflux much better for wild-type than mutant ABCG2, bound similarly to purified ABCG2, while the highly-potent Ko143 bound in the nanomolar range also effective in inhibition of drug transport. All results indicate that the role of the arginine-482 mutation on substrate drug transport and inhibitor efficiency is not mediated by changes in drug binding. Received 10 April 2006; received after revision 22 May 2006; accepted 12 June 2006 A. Pozza and J. M. Perez-Victoria contributed equally to this work     

The kinetics of estrogen binding to rat α-fetoprotein

Summary Fluid obtained from rat fetuses was utilized to characterize the affinity, number of binding sites, and the association and dissociation rate kinetics of the binding of estradiol and estrone to AFP. Statistical analysis demonstrated no differences when the values for the AFP-estradiol interaction were compared with those obtained for the AFP-estrone interaction. These data demonstrate that rat AFP specifically binds estradiol and estrone with a high capacity, high affinity, and similar binding kinetics.This work was supported by NSF grant PCM-8109847 and by a Grant-in-Aid of Research from Sigma Xi, The Scientific Research Society.