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Biomedical subjects

C Montecucco

Publications and source records attributed to C Montecucco.

At least 217 records · Page 12Linked to original sources
PubMed1987

Diphtheria toxin and its mutant crm 197 differ in their interaction with lipids.

The interaction of diphtheria toxin and its enzymatically deficient mutants crm 176 and crm 197 with liposomes has been studied by turbidity measurement and hydrophobic photolabelling with photoactivatable phosphatidylcholines. Diphtheria toxin and crm 176 at neutral pH bind to the surface of lipid bilayers while crm 197 also appears to interact with the fatty acid chains of phospholipids. All proteins undergo a change in conformation over the same range of acidic pH and become able to insert in the lipid bilayer. The tighter lipid interaction of crm 197 may account for its higher cell association constant. The possibility is discussed that the binding of diphtheria toxin to cells is mediated by both a protein receptor and an interaction with the head group of phospholipids.

Diphtheria Toxin↗
PubMed1987

Photolabeling of the integral proteins of skeletal muscle sarcoplasmic reticulum: comparison of junctional and nonjunctional membrane fractions.

Rabbit skeletal muscle sarcoplasmic reticulum (SR) was fractionated by isopycnic density gradient centrifugation into longitudinal tubules (LSR) and terminal cisternae (TC). Junctional face membranes (JFM) were obtained by Triton X-100 treatment of the TC fraction (Costello, B., Chadwick, C., Saito, A., Chu, A., Maurer, A. and Fleischer, S. (1986) J. Cell Biol. 103, 741-753). Photoactivatable phospholipid analogs were introduced into LSR, TC, and JFM fractions to specifically label integral membrane proteins. Remarkably different labeling patterns were observed. Proteins of the following Mr were labeled and identified in the junctional sarcoplasmic reticulum (JFM): 350,000, 325,000, 80,000, 49,000, 37,000, 32,000, 30,000, and 6000. Polypeptides of Mr 105,000 (Ca2+-dependent ATPase), 77,000, 55,000, 41,000, 22,000, and 9000 (proteolipid) were labeled and found to be selectively localized in the nonjunctional sarcoplasmic reticulum (LSR). Calsequestrin, a key protein responsible for Ca2+ storage within the SR lumen, was never labeled, whether 1 mM CaCl2 was present or absent, and is termed a nonintegral membrane protein.

Affinity Labels↗
PubMed1987

Isolation and characterization of cytochrome c oxidase from bird and fish heart mitochondria.

1. Several bird and fish heart mitochondrial cytochrome c oxidases have been isolated with a rapid and simple method involving hydrophobic and affinity chromatography. 2. Their spectrophotometric and kinetic properties are very similar to those of the mammalian enzymes. 3. These oxidases show a polypeptide composition simpler than the mammalian enzymes being composed of 9-10, instead of 13, different polypeptides. 4. These data suggest that the complexity of the mitochondrial heart oxidase increases with the stage of evolution.

Animals↗
PubMed1986

High-degree abnormalities of nuclear DNA distribution in SIg negative acute lymphoblastic leukemia with vacuolated blasts.

Among 65 consecutive patients with untreated adult acute lymphoblastic leukemia (ALL), six had peculiar clinical and cytologic features. Bone marrow and circulating blasts were large and showed prominent vacuolization, with variable degree of granular periodic acid-Schiff positivity in the cytoplasm. All patients had marked spleen and liver enlargement with cholestasis and without either lymph node or central nervous system disease. They achieved complete response to chemotherapy, and their median survival was longer than 12 months. Blast cells were studied for immunologic markers, propidium iodide flow cytofluorometric DNA content, and in vitro tritiated thymidine labeling index (3H-TdR LI). These showed a "null" phenotype in two cases, contained intracytoplasmic immunoglobulins (Cy mu) in two and were positive for CALL and Ia antigens in two others (one of which also contained Cy mu). Surface immunoglobulins (SIg) were found on less than 12% of cells, and T-cell markers were absent. In five cases, one or two stem lines with highly abnormal modal DNA content were found to coexist with diploid blasts. The 3H-TdR LI was high (8.5%-15.5%). Abnormal cell lines, after being cleared by chemotherapy, were found again at relapse. Additional cell lines appeared during the course of the disease in one patient, having different sensitivity to cytostatics. These cases of ALL apparently derive from B-cells not yet expressing SIg and have an exceptionally high incidence of aneuploid cell lines.

Adolescent↗
PubMed1986

A pH-induced increase in hydrophobicity as a possible step in the penetration of colicin E3 through bacterial membranes.

Exposure to low pH triggers an increase in the hydrophobicity of the colicin E3 molecule. Using a [3H] Triton X-100 binding assay we have shown that the amount of detergent (at supramicellar concentrations) associated with colicin E3 increased dramatically at pH 3.8 and below. Interaction of colicin E3 with asolectin vesicles was monitored by following its cross-linking with two different photoactivatable radioactive phospholipid analogues. At neutral pH colicin E3 was cross-linked with the phospholipid probing the membrane surface whereas at pH 4.5 and below, the bacteriocin reacted with the phospholipid probing the hydrophobic core of the bilayer. With the use of phase partitioning of proteins in Triton X-114 it was shown that at acidic pH whole colicin E3 and its immunity protein segregated in the detergent phase. After trypsin digestion of the colicin-immunity complex, the N-terminal portion of E3 (T1) and the immunity partitioned in the detergent phase at low pH. In contrast, the enzymic domain of the colicin (T2) remained in the aqueous phase and was recovered in a highly active form as a consequence of its dissociation from the immunity protein. These results are discussed in relation to the mechanism of entry of colicin E3 into bacterial cells.

Carbon Radioisotopes↗
PubMed1986

1-Palmitoyl-2-(p-benzoyl)benzoyl phosphatidylcholine, a photoactive phospholipid for the labelling of membrane components.

The preparation and use of a new photoactivatable phosphatidylcholine derivative [1-palmitoyl-2-(p-benzoyl)benzoyl phosphatidylcholine] is described. The reagent is shown to be effective in labelling different membrane proteins under irradiation conditions that preserve their enzymic activity. The properties of this new phospholipid analogue suggest that it can be useful in studying several aspects of membrane structure and function.

Adenosine Triphosphatases↗
PubMed1986

Tetanus toxin is labeled with photoactivatable phospholipids at low pH.

The mechanism of cell penetration by tetanus toxin is unknown; it has been suggested that the toxin may penetrate into the lipid bilayer from a low-pH vesicular compartment. In this work, the interaction of tetanus toxin with liposomal model membranes has been studied by following its photoinduced cross-linking with either a nitrene or a carbene photolytically generated from corresponding light-sensitive phosphatidylcholine analogues. The toxin was labeled only at pHs lower than 5.5. The low pH acquired hydrophobicity of tetanus toxin appears to be confined to its light chain and to the 45-kDa NH2-terminal fragment of the heavy chain. Negatively charged lipids promote the interaction of this toxin with the hydrocarbon chain of phospholipids. The relevance of the present findings to the possible mechanism of nerve cell penetration by tetanus toxin is discussed.

Carbon Radioisotopes↗
PubMed1986

ATP binding to bovine heart cytochrome c oxidase. A photoaffinity labelling study.

ATP influences the kinetic properties of cytochrome c oxidase. A photoactivatable radioactive ATP analogue was used to localize the nucleotide-binding site on the bovine heart enzyme. Subunits IV and VIII were specifically labelled, suggesting that these two nuclear-coded polypeptides may play a regulatory role on the oxidase functions.

Adenosine Triphosphate↗
PubMed1986

Hydrophobic photolabelling of pertussis toxin subunits interacting with lipids.

The hydrophobic surfaces presumably involved in the membrane interaction of pertussis toxin have been mapped by a new detergent-binding assay. This is based on the interdispersion among detergent micelles of trace amounts of radioactive photoreactive phospholipid analogues, able to cross-link to the protein thereby labelling its detergent-binding domains. The assay has proven to be very sensitive. Subunits B1, B2 and B3 of pertussis toxin were found to interact with the lipid micelles suggesting that they may be involved in the membrane penetration step of the intoxication process.

Detergents↗
PubMed1986

Peptichemio, vincristine and prednisone versus melphalan and prednisone as induction therapy in multiple myeloma.

Seventy-five patients with previously untreated multiple myeloma were randomly treated with the association of Peptichemio, Vincristine and prednisone (PTC-VCR-P) or of melphalan and P (MPH-P) for first induction therapy. After induction, all responsive patients received MPH and P until relapse, while unresponsive patients received it until unequivocal evidence of disease progression was observed. A second induction therapy with PTC-VCR-P was then administered, except to patients resistant to this association at first induction (who received combination chemotherapy which included cyclophosphamide and adriamycin). The response rate was 58% in the PTC-VCR-P and 41% in the MPH-P group (P greater than 0.05). The PTC-VCR-P responsive patients experienced a median duration of response shorter than MPH-P responsive patients (20.3 vs 39.7, P = 0.041). Median survival from the start of treatment was 26.2 months in the PTC-VCR-P and 54.1 months in the MPH-P group of patients (P = 0.039). Stage I and II myelomas had the same response rate to PTC-VCR-P and to MPH-P, but their survival was shorter on PTC-VCR-P than on MPH-P (P = 0.014). Stage III myelomas responded more frequently to PTC-VCR-P than to MPH-P (P less than 0.02) and there was a trend to survive longer on PTC-VCR-P than on MPH-P (22.0 vs 12.5 months, P greater than 0.05).

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1986

Comparative biochemistry of the ubiquinol-cytochrome c oxidoreductase (EC 1.10.2.2) isolated from different heart mitochondria.

The ubiquinol-cytochrome c oxidoreductase (bc1 complex, EC 1.10.2.2) has been isolated from the heart mitochondria of beef, chicken, turkey, duck and tuna with an identical procedure. The polypeptide composition of the different complexes, compared using SDS-polyacrylamide gel electrophoresis, shows that the three subunits carrying the prosthetic groups of the enzyme are highly conserved in all species. Also the large subunits I and II (core proteins) and band VI appear to be conserved in structure, while subunits VII and VIIa show a most remarkable structural variation in the various complexes. The steady-state ubiquinol-cytochrome c reductase analysis of the active enzymes indicates that all the bc1 complexes follow essentially a ping-pong mechanism, with the cytochrome c substrate displaying a partial competitive inhibition vs the ubiquinol substrate. The cytochrome c specificity of the reductase activity clearly is different in the various bc1 complexes, whereas the quinol specificity appears to be identical in all the enzymes.

Animals↗
PubMed1986

Complete remission in plasma cell leukaemia.

Two patients with primary plasma cell leukaemia who achieved complete remission are reported. They were treated with induction therapy consisting of a multipeptide derivative of sarcolysin, Peptichemio, given intravenously, combined with vincristine and/or prednisone, followed by conventional melphalan-prednisone therapy. 5-7 months following the beginning of therapy, both patients attained a complete remission which lasted 23 and 6 months; second remission was not achieved. Survival from starting therapy was 57 and 16 months respectively. These cases indicate that intravenous alkylating agents can induce a complete remission in plasma cell leukaemia similar to that achieved in other acute leukaemias.

Aged↗
PubMed1986

Adult acute non-lymphoblastic leukaemia: reliability and prognostic significance of pretreatment bone marrow S-phase size determined by flow cytofluorometry.

46 adult patients with non-lymphoblastic acute leukaemia (AnLL) had pretreatment proliferative activity of bone marrow (BM) blasts determined simultaneously with propidium iodide DNA flow cytofluorometry (as the percentage of cells with DNA content intermediate between the diploid and the tetraploid values, 2n-4n cell %) and in vitro tritiated thymidine cytoautoradiography (as the labelling index, LI). They were then treated with the same chemotherapy regimen, including 2-3 courses of sequential vincristine, arabinosylcytosine and adriamycin, for response induction, and monthly courses of different cytostatics for maintenance. Median 2n-4n cell % was 9.9 and median LI was 5.9. A close linear relationship (r = 0.913, p less than 0.001) between the two parameters was found. Patients having a 2n-4n cell % greater than 15.6 responded more often to chemotherapy than patients having a lower percentage (p less than 0.05) but also experienced a shorter duration of first response (p less than 0.05). Overall survival was longer in patients with 2n-4n cell % greater than 15.6 (p less than 0.02). Multiple regression analysis indicates that 2n-4n cell % is a statistically significant (p less than 0.05) independent factor correlating with duration of response in adult AnLL.

Acute Disease↗
PubMed1986

Analysis of human myeloma cell population kinetics.

Pretreatment plasma cell 3H-thymidine labeling index (LI) was related to chemotherapy response in 37 multiple myeloma patients treated with alkylating agents. Response rate did not significantly differ in patients with high (greater than 3%), intermediate (2-3%) and low (less than 2%) LI values, and survival duration was significantly longer in patients with low LI irrespective of response to chemotherapy. Median response duration was distinctly shorter in patients with high and intermediate values (3 and 11 months, respectively) than in those with low values (30 months; p less than 0.01). LI at relapse was similar to that found at presentation in 5 out of 10 patients studied but was sharply increased in the other 5 who had an aggressive clinical course following initial response. Four of these latters showed a clonal evolution of bone marrow plasma cells, detected by changes in cell ploidy at flow-cytometric analysis.

Cell Cycle↗
PubMed1985

pH-dependence of the phospholipid interaction of diphtheria-toxin fragments.

Photoreactive phospholipids have been used to probe the lipid interaction of diphtheria toxin. Low pH values induce the membrane insertion of both the binding and enzymic fragments of the toxin. The efficiency of this process is much higher with asolectin than with egg lecithin (phosphatidylcholine)/cholesterol liposomes. The low-pH-induced interaction of the toxin fragments with the membrane hydrocarbon phase is more evident for the enzymic A-chain than for the binding B-chain, and it is fully reversed by returning the pH to neutrality.

Diphtheria Toxin↗