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Biomedical subjects

I Pecht

Publications and source records attributed to I Pecht.

215 records · Page 12Linked to original sources
PubMed1972

Electron transfer to ferricytochrome c: reaction with hydrated electrons and conformational transitions involved.

The reaction of horse-heart cytochrome c with hydrated electrons has been studied by the pulseradiolysis technique. In neutral solution, the ferriheme group was reduced in a bimolecular reaction that takes place at a rate equal to that of the decay of the e(-) (aq), and approaches the diffusion-controlled limit. This reduction is assigned mainly to a direct reaction, proceeding via the exposed edge of the porphyrin projecting into the cytochrome c crevice. The reaction absorption spectrum observed 20 musec after an electron pulse was very similar, yet blue-shifted relative to, the difference spectrum between the reduced and oxidized forms of cytochrome c. However, this shift vanishes in a slow monomolecular reaction, which seems to reflect the conformational relaxation of the protein to the final equilibrium state of its reduced form. In alkaline solutions, the transition of cytochrome c molecules into an irreducible conformation causes a proportionate decrease in the amount of ferricytochrome c reduced in the direct reaction. The rate of conformational transition of the protein into the reactive form is now the limiting step for a substantial part of the reduction that takes place via this slow monomolecular reaction. Pseudomonas cytochrome c 551 which, in contrast to horse-heart cytochrome c, is a negatively charged protein at neutral pH reacts with e(-) (aq) at a rate lower than does the horse-heart protein. The reduction of the heme group follows that of the e(-) (aq) decay with a small, yet significant, delay.

Animals↗
PubMedDate not supplied

Basophil variants with impaired cromoglycate binding do not respond to an immunological degranulation stimulus.

Cromoglycate, which inhibits IgE-mediated degranulation of mast cells and a basophilic rat tumour cell line (RBL-2H3) (ref.3), is a drug widely used in the treatment of allergic asthma. We have demonstrated the presence of specific binding sites for that drug on the membranes of basophils and mast cells and more recently, we have succeeded in isolating the cromoglycate-binding protein from the membranes of RBL-2H3 cells. These findings together with the chelation by cromoglycate of alkaline-earth ions in low polarity medium, suggest that the binding site of the drug may either be part, or closely related to the calcium gate. To further investigate this, we have selected variants of the RBL-2H3 cell line that are defective in cromoglycate binding but bind IgE normally. In these variants, which have similar histamine content to the parental cells, IgE-mediated challenge did not lead to Ca2+ influx, degranulation and histamine release. In contrast, these cells were able to release histamine on exposure to the Ca2+ ionophore A23187, indicating that the degranulation mechanism distal to the Ca2+ gating step is unaffected. Taken together, these findings suggest that cromoglycate-binding protein has a role in the transmembrane calcium influx which induces degranulation.

Animals↗
PubMedDate not supplied

Protein kinase C regulation of the receptor-coupled calcium signal in histamine-secreting rat basophilic leukaemia cells.

It has been proposed that protein kinase C mediates cellular responses evoked by external stimuli, leading to alterations in internal free calcium concentrations. We have shown previously that histamine-secreting rat basophilic leukaemia cells (RBL-2H3), which degranulate on aggregation of the receptors for immunoglobulin IgE, contain a Ca2+- and phospholipid-dependent protein kinase (kinase C). The partially purified enzyme is activated directly by the tumour-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA). In intact RBL cells, TPA potentiates histamine release induced by the Ca2+-ionophore A23187 (similar to the synergy reported for platelets, neutrophils and rat peritoneal mast cells). Although TPA at concentrations below 15 nM synergizes with the antigen, higher TPA concentrations inhibit secretion. This selective inhibition suggested that kinase C is involved in both the activation and termination of the secretory process. To examine this possibility, we have determined the effect of TPA on changes in free cytosolic Ca2+ concentration during antigen-induced release. We report here that TPA completely blocks the increase in Ca2+ concentration induced by antigen. Our results strongly suggest that protein kinase C is involved in the regulation of receptor-dependent Ca2+ signalling.

Basophils↗
PubMedDate not supplied

A new member of the C-type lectin family is a modulator of the mast cell secretory response.

A glycoprotein identified on RBL-2H3 cells as capable of inhibiting the secretory response induced by the type I Fc epsilon receptor was named mast-cell-function-associated antigen (MAFA). The amino acid sequence deduced from the cloned full-length cDNA has now shown that the MAFA has marked sequence homology with several members of the C-type (calcium-dependent) animal lectin family. The high conservation of cysteinyl residues suggests an important role for intrachain disulfide bonds in attaining its structure and biological activity. We further show that MAFA clustering by monoclonal antibody G63 also inhibits the de novo synthesis and secretion of interleukin-6 induced by the Fc epsilon RI stimulus. Though no ligand has yet been identified for the MAFA, experiments using antisense oligonucleotides suggest that this novel lectin may have a role in cell adhesion in addition to its immunomodulatory capacity.

Amino Acid Sequence↗