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Biomedical subjects

R Adar

Publications and source records attributed to R Adar.

At least 37 records · Page 2Linked to original sources
PubMed1996

Mutational studies of the amino acid residues in the combining site of Erythrina corallodendron lectin.

High-resolution X-ray crystallography of the complex of the Gal/GalNAc-specific Erythrina corallodendron lectin with lactose identified the amino acid side chains that form contacts with the galactose moiety of the disaccharide. The contribution of these amino acids to the binding of different monosaccharides and oligosaccharides by the lectin was examined by site-directed mutagenesis. Replacement of Phe131, on which the galactose is stacked, by tyrosine, gave a mutant with the same hemagglutinating activity and carbohydrate specificity as the parent lectin, but replacement by alanine or valine resulted in loss of activity. Mutations of Ala88, Asp89, and Asn133 produced mutants that were also inactive whereas those of the other combining site residues, Tyr106, Ala218, and Gln219, were biologically active. None of the active mutants interacted with mannose or glucose. Thus, contrary to an earlier assumption. Ala218 is not responsible for the inability of E. corallodendron lectin to bind these sugars. Our findings also demonstrate that Gln219 is not involved in galactose binding in solution, even though this is implicated by the crystal data. Instead, our data suggest that Gln219 assists in the ligation of N-acetyllactosamine to the lectin, by interacting with the acetamide group of the disaccharide. Comparison with other legume lectins specific for mannose/glucose, galactose, N-acetylgalactosamine, L-fucose or N-acetylglucosamine, shows that only three of the combining site residues of E. corallodendron lectin occupy invariant positions both in their primary and tertiary structures. These residues are an aspartic acid and an asparagine corresponding to positions 89 and 133, respectively, in E. corallodendron lectin, and an aromatic residue, either phenylalanine (as Phe131 in this lectin), tyrosine or tryptophan. We therefore postulate that these three residues are essential for ligand binding by all such lectins, irrespective of their specificity.

Acetylgalactosamine↗
PubMed1996

Vascular complications of total abdominal perfusion and aortic stop-flow infusion.

During a 2 year period (1992-1993), 149 patients with advanced abdominal cancer underwent total abdominal ischemic perfusion (TAP) and stop-flow infusion (SFI) 159 times in an attempt to achieve palliation. These procedures and aortic stop-flow infusion require insertion of balloon catheters into the abdominal aorta and inferior vena cava by a transfemoral approach. Flow is arrested for 15 minutes, during which time chemotherapeutic agents are infused into the aorta, distal to the balloon occlusion. Femoral access is by a surgical incision. The passage of the catheters is guided by fluoroscopy. Some tumor response was observed in 35% of the patients. Ten patients had major vascular complications; two iliac artery aneurysms were lacerated and required emergency repair. There were two femoral artery false aneurysms that required surgical correction, one early and one late. Aortic dissection was detected in four patients, but these did not require surgical intervention. Two patients had thrombosis distal to the occluded vessel, both required surgical intervention. To reduce the incidence of these vascular complications we recommend: (1) a clinical and vascular laboratory evaluation before the procedure, and (2) angiography of normal flow in patients with underlying vascular disease.

Abdominal Neoplasms↗
PubMed1996

A point scoring system for the clinical diagnosis of Buerger's disease.

INTRODUCTION: The literature on thromboangiitis obliterans (TAO, Buerger's disease) suffers from the lack of a unified method of establishing the diagnosis of the disease. The aim of this study was to test the application of a point scoring system (PSS) in the diagnosis of TAO. METHOD: Points are awarded for young age at onset, foot claudication, upper extremity involvement, superficial vein thrombosis and vasospastic phenomena. Atypical features detract points, and the resultant score classifies the diagnosis of TAO as being of low, medium or high probability. One hundred and seven patients diagnosed and classified according to our previous accepted criteria for TAO, were independently reclassified by the PSS, and the results of the two diagnostic processes were compared. RESULTS: Of the 107 patients diagnosed by our old criteria (OC) the diagnosis of TAO was rejected by the PSS in 20 patients. Of the remaining 87 patients the degree of certainty in the diagnosis (PSS vs. OC) was lower in 31 equal in 47 and higher only in nine. CONCLUSION: The proposed PSS is more discriminating than our OC. If this type of scoring system can be agreed upon the certainty in the diagnosis of TAO would be increased. Data collection would be improved, and the cause of clinical and basic research would be advanced.

Adult↗
PubMed1996

Exploring cell death mechanisms by analyzing signaling cascades of the TNF/NGF receptor family.

The ability of ligands of the tumor necrosis factor (TNF) family to induce death of cells independently of new protein synthesis provides a unique approach to molecular analysis of programmed cell death mechanisms. Sequential analysis of the protein-protein interactions by which these receptors signal, allows identification of specific molecules that participate in the cell death process and unequivocal definition of cause-effect relationships between them. Several receptors of this family, with structurally unrelated intracellular domains, have the ability to trigger cell death. some intracellular proteins that bind to the receptors and participate in the induction of their effects have been identified. Association of the Fas/APO1-interacting protein MORT1/FADD with the p55 TNF receptor-interacting protein TRADD, and the association of both MORT1/FADD and TRADD with a third protein, RIP, provide potential cross-talk mechanisms between Fas/APO1 and the p55 TNF receptor. TRAF2, a cytoplasmic protein that binds to the p75 TNF receptor, as well as to several other receptors of the TNF/NGF family, also binds to TRADD, thus further extending the range of receptors of this family that can share common signaling mechanisms. The N-terminal part of MORT1/FADD binds to a protease of the CED3/ICE family, MACH alpha. Activation of MACH alpha by the TNF/NGF receptors appears to be the most upstream enzymatic activity in the cascade of signaling for cell death.

Adaptor Proteins, Signal Transducing↗
PubMed1995

Close similarity among streptavidin-like, biotin-binding proteins from Streptomyces.

Two strains of Streptomyces venezuelae were found to produce high-affinity, biotin-binding proteins, termed streptavidin v1 and v2, respectively. Both proteins were isolated to purity, and their corresponding genes were cloned and sequenced. Compared to streptavidin from S. avidinii, streptavidin v1 had only a single amino acid substitution and streptavidin v2 showed 9 such differences. The substitutions were remarkably conservative, none of which affected the amino acid residues known to be important to the biotin-binding properties or to the structure of the tetrameric protein. The results also indicate that the biosynthesis of such biotin-binding proteins is not simply a curious anomaly in a single species of Streptomyces. It is suggested that the classification of S. avidinii as a unique species should be reconsidered. The occurrence of these proteins appears to be linked to the production of an unusual synergistic antibiotic complex.

Amino Acid Sequence↗
PubMed1995

C-terminal post-translational proteolysis of plant lectins and their recombinant forms expressed in Escherichia coli. Characterization of "ragged ends" by mass spectrometry.

Electrospray mass spectrometry was used to accurately measure the molecular masses of single chain lectins from legume seeds and also of three recombinant lectins, expressed in Escherichia coli. The five single chain lectins, Erythrina corallodendron lectin, soybean and peanut agglutinins, Dolichos biflorus lectin, and Phaseolus vulgaris hemagglutinin E, all showed evidence of C-terminal proteolytic processing, in some cases to "ragged" ends, when their masses were compared to those expected from their cDNA sequences and their known carbohydrate chains. Recombinant forms of the lectins from E. corallodendron, soybean, and peanut also showed C-terminal trimming, but not to the same points as the natural forms. Discrepancies between the protein and cDNA sequences of the E. corallodendron lectin were resolved by combined liquid chromatography-mass spectrometry peptide mapping and protein sequencing experiments, and the presence of a second glycosylation site was demonstrated. Our data show that all of these lectins undergo C-terminal proteolytic processing of a readily attacked peptide segment. This trimming is frequently imprecise, and the resulting heterogeneity may be a major contributor to the appearance of isolectin forms of these proteins.

Amino Acid Sequence↗
PubMed1995

Expression of fibrinolytic genes in atherosclerotic abdominal aortic aneurysm wall. A possible mechanism for aneurysm expansion.

Expansion of atherosclerotic abdominal aortic aneurysm (AAA) has been attributed to remodeling of the extracellular matrix by active proteolysis. We used in situ hybridization to analyze the expression of fibrinolytic genes in aneurysm wall from eight AAA patients. All specimens exhibited specific areas of inflammatory infiltrates with macrophage-like cells expressing urokinase-type plasminogen activator (u-PA) and tissue-type PA (t-PA) mRNA. Type 1 PA inhibitor (PAI-1) mRNA was expressed at the base of the necrotic atheroma of all specimens and also within some of the inflammatory infiltrates where it frequently colocalized in regions containing u-PA and t-PA mRNA expressing cells. However, in these areas, the cellular distribution of the transcripts for t-PA and u-PA extended far beyond the areas of PAI-1 expression. These observations suggest a local ongoing proteolytic process, one which is only partially counteracted by the more restricted expression of PAI-1 mRNA. An abundance of capillaries was also obvious in all inflammatory infiltrates and may reflect local angiogenesis in response to active pericellular fibrinolysis. The increased fibrinolytic capacity in AAA wall may promote angiogenesis and contribute to local proteolytic degradation of the aortic wall leading to physical weakening and active expansion of the aneurysm.

Adult↗
PubMed1994

Modulation of the fibrinolytic system by major peripheral ischemia.

PURPOSE: A rat model was developed to investigate the effects of acute peripheral ischemia on the components of the fibrinolytic system. METHODS: Laparotomy was performed and ischemia was introduced by total aortic clamping at a subrenal position. Control animals underwent sham laparotomy alone. Plasma and tissue samples were collected for analysis at 30, 60, 90, and 120 minutes after operation. RESULTS: Functional assays of rat plasma revealed a dramatic and transient increase in tissue-type plasminogen activator (tPA) activity within 30 minutes of the onset of ischemia. A simultaneous decline in plasminogen activator inhibitor activity was observed. Immunohistochemical analysis suggested this initial increase in tPA activity resulted primarily from the release of stored tPA from ischemic vascular tissues. Northern blot analysis revealed that both tPA and plasminogen activator inhibitor-1 messenger RNA levels were elevated at 60 to 120 minutes in well-perfused tissues distant from the ischemic insult. CONCLUSIONS: Collectively, these data demonstrate that acute peripheral ischemia results in a rapid and transient increase in plasma fibrinolytic activity, concomitant with the early release of stored tPA from ischemic vascular tissues. In addition, peripheral ischemia appears to stimulate both tPA and plasminogen activator inhibitor-1 gene expression in well-perfused tissues at later time points, consistent with the existence of humoral mediators.

Adipose Tissue↗
PubMed1994

Surgical treatment of palmar hyperhidrosis before thoracoscopy: experience with 475 patients.

Between the years 1968-1992, 475 patients underwent simultaneous bilateral upper dorsal sympathectomy by the supraclavicular approach for severe palmar hyperhidrosis. For the purpose of comparing outcomes of the open surgical method with the increasingly used thoracoscopic procedure, we reviewed the clinical data of our patients. The incidence of severe palmar hyperhidrosis in the young population in Israel is 1-2/1,000. Surgical excision of the T2 and T3 ganglia was effective in drying the hands of all patients, who had frozen section confirmation of removal of a ganglion. At follow-up, hyperhidrosis recurred in 5.3% of limbs. Mild transient Horner's syndrome occurred in 12% of procedures, but only in 5 patients was it permanent. The main drawback of the open surgical approach lies in the postoperative complications. The effectiveness of the thoracoscopic approach will be judged by immediate and late results, and by the expected reduction in postoperative morbidity.

Hand↗
PubMed1994

Carotid endarterectomy for symptom-free stenosis: the patient's point of view.

Determining the indication for carotid endarterectomy in a given patient with a totally symptom-free internal carotid artery stenosis depends on objectively assessing the risk of non-operative versus operative treatment, as well as on the patient's attitude towards immediate versus long-term risk. In order to study patients' attitudes and preferences, four groups of subjects were asked to complete two questionnaires: the first examined people's perceived deterioration in their quality of life if they sustained a stroke of a defined severity. The second checked the attitude towards risk of operative versus non-operative treatment at different levels of an expected annual stroke rate without surgery. This preliminary study showed the feasibility of studying these subjective elements, and also pointed to problem areas in this type of research. The existence of individual as well as group differences in patients' values and preferences should be considered in the decision-making process.

Adult↗
PubMed1993

Modification by site-directed mutagenesis of the specificity of Erythrina corallodendron lectin for galactose derivatives with bulky substituents at C-2.

Examination of the three-dimensional structure of Erythrina corallodendron lectin (ECorL) in complex with a ligand (lactose), the first of its kind for a Gal/GalNAc-specific lectin [(1991) Science 254, 862-866], revealed the presence of a hydrophobic cavity, surrounded by Tyr108 and Pro134-Trp135, which can accommodate bulky substituents such as acetamido or dansylamido (NDns) at C-2 of the lectin-bound galactose. Comparison of the primary sequence of ECorL with that of soybean agglutinin, specific for galactose and its C-2 substituted derivatives, and of peanut agglutinin, specific for galactose only, showed that in soybean agglutinin, Tyr108 is retained, and Pro134-Trp135 is replaced by Ser-Trp, whereas in peanut agglutinin, the former residue is replaced by Thr and the dipeptide by Ser-Glu- Tyr-Asn. Three mutants of ECorL were therefore constructed: L2, in which Pro134-Trp135 was replaced by Ser-Glu-Tyr-Asn; Y108T, in which Tyr108 was replaced by Thr and the double mutant L2; Y108T. They were expressed in Escherichia coli, as done for recombinant ECorL [(1992) Eur. J. Biochem. 205, 575-581]. The mutants had the same hemagglutinating activity as native or rECorL. Their specificity for galactose, GalNAc and Me beta GalNDns was examined by inhibition of hemagglutination and of the binding of the lectin to immobilized asialofetuin; in addition, their association constants with Me alpha GalNDns and Me beta GalNDns were measured by spectrofluorimetric titration. The results showed that Y108T had essentially similar specificity as the native and recombinant lectins. The affinity of L2 and L2;Y108T for galactose was also the same as ECorL, but they had a lower affinity for GalNAc and markedly diminished affinity for the dansyl sugars (up to 43 times, or 2 kcal, less). This appears to be largely due to steric hindrance by the two additional amino acids present in the cavity region in these mutants. Our findings also provide an explanation for the inability of PNA to accommodate C-2-substituted galactose derivatives at its primary subsite.

Amino Acid Sequence↗
PubMed1992

Cloning, sequence analysis and expression in Escherichia coli of the cDNA encoding a precursor of peanut agglutinin.

The cDNA coding for pre-peanut agglutinin (PNA) was isolated from a bacterial expression library. It codes for a polypeptide of 273 amino acids composed of a hydrophobic signal peptide of 23 amino acids and a mature protein of 250 amino acids. The sequence of the latter is identical to that of native PNA, determined very recently by conventional methods, except that it contains 14 additional amino acids at the C-terminus. Bacterial cells harboring a plasmid with the prePNA-cDNA, produced two PNA cross-reacting proteins: one migrated on SDS-PAGE identically with the native lectin (apparent mol. wt. 31 kDa); the other, at 35 kDa, was a beta-galactosidase pre-PNA fusion protein. The former protein possessed an N-terminal sequence identical to that of the mature, native PNA, suggesting that it was processed from the 35 kDa prePNA precursor. Only the 31 kDa protein was exported into the bacterial periplasmic space, and had the ability to bind to galactose-Sepharose. The isolated processed protein had the same hemagglutinating activity as the native lectin, when assayed with sialidase-treated human erythrocytes. Like the native lectin, it did not agglutinate the untreated cells, was not inhibited by N-acetylgalactosamine, and was inhibited by Gal beta 1----3GalNAc 30-times more strongly than by galactose.

Amino Acid Sequence↗