Showing papers on "Polyamine binding published in 2006"

The Polyamine Binding Site in Inward Rectifier K+ Channels

Abstract: Strongly inwardly rectifying potassium channels exhibit potent and steeply voltage-dependent block by intracellular polyamines. To locate the polyamine binding site, we have examined the effects of polyamine blockade on the rate of MTSEA modification of cysteine residues strategically substituted in the pore of a strongly rectifying Kir channel (Kir6.2[N160D]). Spermine only protected cysteines substituted at a deep location in the pore, between the “rectification controller” residue (N160D in Kir6.2, D172 in Kir2.1) and the selectivity filter, against MTSEA modification. In contrast, blockade with a longer synthetic polyamine (CGC-11179) also protected cysteines substituted at sites closer to the cytoplasmic entrance of the channel. Modification of a cysteine at the entrance to the inner cavity (169C) was unaffected by either spermine or CGC-11179, and spermine was clearly “locked” into the inner cavity (i.e., exhibited a dramatically slower exit rate) following modification of this residue. These data provide physical constraints on the spermine binding site, demonstrating that spermine stably binds at a deep site beyond the “rectification controller” residue, near the extracellular entrance to the channel.

A new approach to DNA bending by polyamines and its implication in DNA condensation.

Abstract: Polyamines are known to induce dynamical bending of DNA molecules. This mechanism is very important since many DNA binding proteins (DNAse, transcription factor, etc.) exert their action by their ability to bend DNA. We propose an analytical model which describes the dynamical bending of DNA by polyamine ions in highly diluted DNA solutions. The bending probability depends on the entropy loss of polyamines due to their localization. This localization is facilitated by the electrostatic repulsion between multivalent counterions condensed on DNA, which reduces the entropy loss in counterion localization. Therefore DNA bending by polyamines depends on the competition between monovalent counterions and polyamines. We find that the bending probability is weak for a low binding ratio of polyamines (i.e. number of bound polyamines per base pair), whereas a high bending probability can be reached at large polyamine binding ratio. In addition, we describe a new mechanism of DNA bending. It occurs with the help of thermal agitation, which initiates the bending and favours the polyamine localization. This model provides further insights into DNA bending by polyamines and its implication in DNA condensation. A qualitative estimation of the DNA bending probability is obtained by measuring the cleavage efficiency of DNA by bleomycin versus spermidine concentration. Indeed, a local helix distortion by polyamines results in an amplification of the double-strand cleavage by bleomycin. The measurement of the bleomycin amplification is performed by analysing images of DNA molecules with atomic force microscope. Some features of the dynamical bending indicate that condensation and bending are interrelated.