Patellamide zinc binding

Table 2. Binding constants of patellamide A and C with copper and zinc...

This would lead us to expect selectivity for copper over zinc as observed as well as preferential binding to copper over nickel. It must be noted that the symmetrical patellamide A is non-selective for Cu, even though the Irving-Williams series would... 

Grondahl et al. found that binding of zinc to ascidiacyclamide (8) was a very slow process. If this is the case with all of the peptides and zinc it would also help explain the observed selectivity for copper over zinc, but when solutions of patellamide C (3) and a mixture of copper and zinc were measured after three months no change had occurred and only the copper complex was in evidence. 

Competition experiments were carried out in a similar manner to those performed using CD. Zinc was added to copper bound patellamide and copper to zinc bound patellamide and the resulting solutions analysed. A mixed solution of copper and zinc was also added to unbound peptide in some cases to ensure that observed selectivity of binding was real and not dependant on which metal was introduced first to the peptide. 

As was seen in the CD competition study (Section 3.4) copper displaced zinc from patellamide C and when a mixture of the two metals was added only copper species were formed. When copper was added to a solution containing predominantly the two zinc complex [PatCfij + Zn2Cl3] at m/z 995 formed by adding a large excess of zinc, this was displaced and the two copper complex [PatCHa + Cu2Cl2] at m/z 957 became the dominant species present. A further competition experiment was carried out with copper bound patellamide C in the presence of acid. The solution of copper and patellamide C was diluted with 2% formic acid and then introduced into the mass spectrometer. No copper species of any kind were detected because the amide NH s were fully protonated at low pH so no binding to Cu was possible. 

These experiments were undertaken in a similar manner to those described by Brady and Sanders for measuring relative binding affinities of metals with steroid derivatives by ESI-MS. Titrations with zinc and copper solutions were carried out on patellamides C and A by sequential addition of 0.25 equivalents of the metal solutions to the peptides (cone. 0.02 mg/mL in MeOH). These titrations were monitored by measuring the formation of the metal species as well as the loss of the uncomplexed peptides, using both the full spectrum and the selected ion mode of the instrument in parallel, as the ionisation efficiency of these species were very different. Once fully complexed zinc species were obtained for each peptide, copper solution was then titrated into these at a rate of 0.25 equivalents to again observe the competition effects. 

The study of these peptides and their copper and zinc complexes by mass spectrometry revealed much information about their composition and binding characteristics. The most interesting results were those from the study of patellamide C and its copper complexes. This revealed the formation of complexes, such as [PatCH2 + 2Cu + (CsHsOs)] which were not detected by CD. This species might be similar to the copper complex of ascidiacyclamide, which was shown by X-ray crystallography to have a bridging carbonate between the two copper atoms. ... 

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