Showing papers on "Polyamine binding published in 1997"
TL;DR: In K+ channels and glutamate receptors, some of the amino acids in the receptor/channel structure that influence to polyamines have been identified, leading to a partial understanding of the effects of polyamines at a molecular level.
241 citations
TL;DR: Observations strongly suggest that this newly defined polyamine-binding domain is involved in the intrasteric regulation of CK2 activity.
95 citations
TL;DR: N-Sulfonyl-polyamines represent a new class of polyamine antagonists with which to study glutamate receptor ion channels and could easily permeate wild-type NR1/NR2A channels, presumably because the head group of N1-OsSpm can pass through the narrowest part of the channel pore.
Abstract: The effects of several N-sulfonyl-polyamines, including N 1 -dansyl-spermine (N 1 -DnsSpm) and N 1 -( n -octanesulfonyl)-spermine (N 1 -OsSpm), were studied at recombinant N -methyl-d-aspartate (NMDA) receptors expressed in Xenopus laevis oocytes. N 1 -DnsSpm and N 1 -OsSpm inhibited NMDA receptors and were ∼1000-fold more potent than spermine in oocytes voltage-clamped at −70 mV. Block by N 1 -DnsSpm and N 1 -OsSpm was strongly voltage dependent, being more pronounced at hyperpolarized membrane potentials. With the Woodhull model of voltage-dependent channel block, the values of K d (0) were 779 μm, 882 μm, and 7.4 mm and those of zδ were 2.58, 2.57, and 1.07 for N 1 -DnsSpm, N 1 -OsSpm, and spermine, respectively. This suggests that an increase in the voltage dependence of block together with an increase in affinity contributes to the increased potencies of N 1 -DnsSpm and N 1 -OsSpm compared with spermine. Sensitivity to N 1 -DnsSpm was reduced by mutation NR1(N616Q) and was increased by mutations NR1(N616G) and NR2A(N615G). The NR1(N616G) and NR2A(N615G) mutations decreased the K d (0) value of N 1 -DnsSpm without affecting zδ, whereas the NR1(N616Q) mutation reduced zδ. These mutations may alter the accessibility of part of the polyamine binding site within the channel pore or directly alter the properties of that site. Block by N 1 -DnsSpm (0.3 μm) was almost complete at −100 mV, and there was no relief of block at extreme negative membrane potentials (−100 to −200 mV) at wild-type NR1/NR2A channels. In contrast, block by N 1 -DnsSpm was partially relieved at extreme negative potentials at receptors containing NR1(N616G) or NR2A(N615G), suggesting that N 1 -DnsSpm can permeate these mutant channels but not wild-type NR1/NR2A channels. This is hypothesized to be due to an increase in the pore size of channels containing NR1(N616G) or NR2A(N615G), which allows passage of the bulky head group of N 1 -DnsSpm. In contrast to N 1 -DnsSpm, N 1 -OsSpm could easily permeate wild-type NR1/NR2A channels, presumably because the head group of N 1 -OsSpm can pass through the narrowest part of the channel pore. N -Sulfonyl-polyamines such as N 1 -DnsSpm and N 1 -OsSpm represent a new class of polyamine antagonists with which to study glutamate receptor ion channels.
68 citations
TL;DR: The elaboration of the first model defining the crucial structural parameters of a polyamine-protein interaction at the molecular level is elaborated, with spermine found to be the most efficient stimulator of the kinase activity and the best CK2 ligand.
Abstract: Protein kinase CK2 is a ubiquitous eukaryotic Ser/Thr kinase whose catalytic activity is enhanced several times by polyamines. We have shown previously that the regulatory beta-subunit of CK2 bears a polyamine binding site located in the region Asp51-Tyr110. In the present study, we have used spermine analogs to investigate the structural requirements of the CK2 polyamine binding site. We have observed a strong correlation between the stimulations of CK2 activity by all tested polyamines and their binding efficiencies to the enzyme. As a result, spermine was found to be the most efficient stimulator of the kinase activity and the best CK2 ligand. The effect of the pH on the stimulation of CK2 activity by spermine strongly suggests the involvement of ionic interactions between the positive charges of spermine and the negative charges of acidic amino acids of the beta-subunit. Using a fusion protein made of MBP and the beta-subunit region encompassing amino acid residues Asp51-Pro110, we have studied the binding of spermine as a function of the ionic strength. We show that this region delineates a functional and autonomous domain containing a binding site involved in the interaction with the four positive charges of spermine. Altogether, these results led to the elaboration of the first model defining the crucial structural parameters of a polyamine-protein interaction at the molecular level.
50 citations
TL;DR: A new and more potent polyamine-conjugated photoprobe, N1-ASA-spermine, has been synthesized and used to develop a method based on detergent lysis for identifying putative cell-surface pbps on solid-tumour cell types that strongly suggests that at least some of the identified pbps may be associated with that function.
Abstract: Polyamine transport is an active process which contributes to the regulation and maintenance of intracellular polyamine pools. Although the biochemical properties of polyamine transport in mammalian cells have been extensively studied, attempts to isolate and characterize the actual protein(s) have met with limited success. As one approach, photoaffinity labelling of cell surface proteins using a polyamine-conjugated photoprobe may lead to the identification of polyamine-binding proteins (pbps) associated with the transport apparatus and/or other regulatory responses. In a previous study [Felschow, MacDiarmid, Bardos, Wu, Woster and Porter (1995) J. Biol. Chem. 270, 28705-28711], we demonstrated that the photoprobes N4-ASA-spermidine and N1-ASA-norspermine [where the ASA (azidosalicylamidoethyl) group represents the photoreactive moiety] competed effectively with polyamines for transport and selectively labelled two major pbps at 118 and 50 kDa on the surface of murine and human leukaemia cells. In the present study, a new and more potent polyamine-conjugated photoprobe, N1-ASA-spermine, has been synthesized and used to develop a method based on detergent lysis for identifying putative cell-surface pbps on solid-tumour cell types. Transport kinetic assays showed that the new photoprobe competed with spermidine uptake with an apparent Ki of 1 microM, a value 20-50-fold lower than those of earlier probes. In L1210 cells, the new probe identified pbp50 and pbp118 thus reaffirming their identity as pbps. Two new bands were also detected. In A549 human lung adenocarcinoma cells, N1-ASA-spermine identified pbps at 39, 62, 73 and 130 kDa, the latter believed to be a size variant of pbp118. The presence of pbp130/118 in two very different cell types suggests the generality of the protein among mammalian cell types as well as its importance for further study. The high affinity of the photoprobe for the polyamine-transport system strongly suggests that at least some of the identified pbps may be associated with that function.
16 citations
Journal Article•
TL;DR: Depletion of intracellular spermidine together with binding to functionally important polyamine binding sites are considered the main features of the compound which cause cytostatic, and at high concentrations, cytotoxic effects.
Abstract: Bis(7-amino-4-azaheptyl)dimethylsilane is a new type of aliphatic polycation with structural features resembling those of spermine. The elongation to a seven-membered carbon chain in which the central CH 2 -group was substituted by (CH3)2Si renders the molecule more lipophilic than spermine. Cells accumulate the compound via the polyamine transport system. Due to suppression of ornithine decarboxylase, de novo putrescine biosynthesis is impaired, and intracellular putrescine and spermidine concentrations are depleted. Depletion of intracellular spermidine together with binding to functionally important polyamine binding sites are considered the main features of the compound which cause cytostatic, and at high concentrations, cytotoxic effects.
12 citations