Selective inactivation of the Arg-Gly-Asp-Ser (RGDS) binding site in von Willebrand factor by site-directed mutagenesis.

In order to assess the requirement for the Arg-Gly-Asp-Ser (RGDS) consensus adhesion sequence in von Willebrand factor (vWF) for vWF binding to platelets and endothelial cells, point mutations were introduced into this sequence by site-directed mutagenesis. A glycine to alanine substitution yielded RADS-vWF, while an aspartate to glutamate substitution resulted in RGES-vWF. Recombinant RADS-vWF and RGES-vWF, purified from transformed Chinese hamster ovary cells, were compared with recombinant wild type vWF (WT-vWF) in functional assays with platelets and human umbilical vein endothelial cells (HU-VECs). High molecular weight RADS-vWF and RGES-vWF multimers inhibited binding of 125I-vWF to a mixture of insolubilized native type I and III collagen and competed effectively with 125I-vWF for binding to formalin-fixed platelets in the presence of ristocetin, indicating functional collagen and platelet glycoprotein Ib binding. However, RADS-vWF and RGES-vWF were unable to displace the binding of 125I-vWF to thrombin or ADP-activated platelets. The attachment of HUVECs to either RADS-vWF or RGES-vWF coated surfaces was reduced and spreading was almost completely inhibited, compared with WT-vWF. We conclude that point mutations of the RGDS sequence in vWF selectively impair binding to platelet glycoprotein IIb/IIIa and the HUVEC vitronectin receptor.     

Apple Proliferation Epidemics Detected in Scab-Resistant Apple Trees

In the Friuli-Venezia Giulia, a region of Italy, where serious epidemics of apple proliferation (AP) are known to occur, varieties resistant to scab (Venturia inaequalis (Cke.) Wint.) are increasingly being used in new orchards. The most important cvs are Florina. Prima and Priscilla. These varieties were cultivated according to organic farming regimes with no insecticides used. The results obtained in two orchards during a 7-year period of investigation indicated that the three varieties resistant to scab are highly susceptible to AP. Florina was most susceptible (high infection rate) to AP while Priscilla was most sensitive (severely affected). The identification of the disease was based on symptom expression, DAPI (4–6-diamidino-2-phenylindole) fluorescence technique, electron microscopy observations and by polymerase chain reaction (PCR). Neither complete spontaneous recovery nor death of AP-infected plants was noticed. The pattern of natural diffusion of AP does not seem to be uniform, the affected trees may be in line or grouped in certain spots of the orchards. This may indicate the activity of a not very mobile vector. It can be concluded that planting of Florina, Prima and Priscilla should be discouraged in areas where AP is a problem, particularly when organic farming regimes are being applied.     

Female sex steroid induced epithelial changes in the gallbladder of the ovariectomized Syrian hamster.

Ovariectomized Syrian hamsters treated by female sex steroids during a 1-month period show gallbladder surface epithelial changes in the fundic area consistent with apical bulging and decapitations of the epithelial cells. These events were detected in the infundibulum and the fundic or body regions of estrogen- and estrogen+progesterone-treated hamsters. In control hamsters, these events were restricted to the region in the vicinity of the bile duct. Following steroid treatment, intraluminal deposits detected resembled Ca-bilirubinate deposits described in previous studies while decapitations are similar to endometrial epithelium changes associated with hormonal physiological changes or treatments. Moreover some small electron-dense deposits are comparable to those found in human cholesterol gallstones. This report indicates that, besides an alteration in bile composition, cell fragments originating from the surface epithelium of the bile duct and/or of the gallbladder mucosal epithelium could participate in gallstone nucleation.     

Properties of carboxypeptidase A-treated chicken gizzard tropomyosin

Chicken gizzard tropomyosin was digested with carboxypeptidase A at the weight ratios of enzyme to substrate 1:200 and 1:50. Removal of about 16 C-terminal amino acid residues per tropomyosin molecule, at lower enzyme concentration, caused reversion of the effect on skeletal actomyosin ATPase activity from activating to inhibiting without an influence on polymerizability and actin-binding ability. Removal of about 26 C-terminal amino acid residues per molecule, at higher enzyme concentration, resulted in loss of polymerizability and actin binding ability. Digestion of gizzard tropomyosin with carboxypeptidase A has no dramatic effect on its binding to troponin T. The results show that not only the existence of head-to-tail overlapping regions but also their length is important for the functional properties of chicken gizzard tropomyosin.     

Effect of the ionic environment and of various neurotransmitters on spontaneous motility in snail intestine (author's transl)]

A spontaneous rhythmic motility of the isolated intestine in snail, Chryptomphalus hortensis, has been registered at 30 degrees C with the aid of an electronic transductor and an appropriate amplification. These slow regular movements are affected by ionic composition changes in the suspension medium. Na+, K+ and Ca++ ions prove to be important in this motility, and the addition of Ba++ markedly stimulates it. ACh produces hypermotility from 1.8 X 10(-11) g/ml. Its effect decreases in the presence of atropine and increases in that of pyridostigmine. The intestine is sensitive to 5-HT from 10(-10) g/ml, which stimulates its activity. The effect of histamine is weak. Low concentrations of adrenaline tend to increase the amplitude, whereas concentrations from 10(-5) g/ml onward produce a cease of motility in relaxation.     

Degradation of commercial detergent products by microbial populations of the Lagos lagoon

The biodegradability potentials of three detergent products with the trade names Omo, Teepol and sodium dodecyl sulfate (SDS) by the native bacteria of the Lagos lagoon was carried out using the lagoon die-away method. Physicochemical parameters of the water samples showed that the lagoon in Apapa was more polluted than at theUniversity of Lagos. In 12 days, approximately 30, 60 and 97% of Omo, Teepol and SDS respectively were degraded. SDS with an alkyl sulfate moiety as surfactant supported the highest growth of the detergent-utilizing organisms, indicating that the components of Omo and Teepol are more resistant to microbial attack. The detergent-utilizing bacteria identified were mainly Gram-negative and of the following genera:Vibrio, Klebsiella, Flavobacterium, Pseudomonas, Escherichia, Enterobacter, Proteus, Shigella andCitrobacter. Vibrio was the most frequently encountered organism whileProteus was the rarest. Results of this investigation had shown that detergents made in Nigeria may still contain components that are recalcitrant to biodegradation.     

Calmodulin-binding proteins also have a calmodulin-like binding site within their structure. The flip-flop model.

The flip-flop model is a mechanistic model proposed to describe how calmodulin activates enzymes. One prediction based upon this model is that calmodulin-activated enzymes would contain a calmodulin-like binding site which, among other attributes, would bind the peptide melittin. Five purified calmodulin-activated enzymes, namely calcineurin, myosin light chain kinase, phosphorylase b kinase, phosphodiesterase, and NAD kinase, were all found to bind biotinylated melittin and to also bind an antimelittin antibody and biotinylated calmodulins. Using gel blots of crude tissue extracts (rat brain and Arabidopsis), most proteins did not bind any of the probes and thus do not have these characteristics. However, among those which bind any of these probes, a strong correlation was found between those proteins which bind biotinylated calmodulins and those which bind melittin and antimelittin. Gel blots of phosphorylase b kinase demonstrate that the alpha, beta, and gamma subunits all bind calmodulin and melittin. A putative calmodulin-like binding site sequence was identified in eight enzymes or subunits which may play an important role in both melittin binding and calmodulin-dependent regulation of these enzymes.