Using diastereopeptides to control metal ion coordination in proteins

Abstract

Here, we report a previously undescribed approach for controlling metal ion coordination geometry in biomolecules by reorientating amino acid side chains through substitution of L-to D-amino acids. These diastereopeptides allow us to manipulate the spatial orientation of amino acid side chains to alter the sterics of metal binding pockets. We have used this approach to design the de novo metallopeptide, Cd (TRIL12L DL16C)

Natural and Artificial Proteins Containing Cadmium

“…This itself was an interesting observation as little to nothing had been reported on the influence on protein structure and stability of D-amino acid substitution into a coiled coil form with L-amino acids. This mixed chirality construct was found to bind 113 Cd(II) with a single 113 Cd NMR chemical shift of 697 ppm [65], which is almost identical to that predicted earlier for 100% CdS 3 (698 ppm) [60]. 111m Cd PAC confirmed the binding site as pure CdS 3 (o o ¼ 0.46 rad/ns) [60].…”

“…TRIL12L D L16C was designed and experimentally shown to bind Cd(II) to Cys as fully bound CdS 3 . The pK a2 associated with Cd(II) binding to the three thiols of Cys as CdS 3 was determined to be 15.1 [65], which is also extremely similar to the value of 15.7 determined for CdS 3 coordination to Pen. These comparisons suggest that the shifts in the pK a2 of binding is due primarily to the resulting Cd(II) coordination geometry, although the alkyl substitution for the penicillamine ligand appears to push up the observed pK a2 by 0.5 to 0.6 units in either coordination geometry.…”

“…113 Cd NMR again supported the selective binding of the first equivalent of 113 Cd(II) to the 4-coordinate site with the appearance of a single resonance at 589 ppm. The second equivalent resulted in the appearance of a second distinct resonance at 690 ppm [65]. These results together with those for GRANDL16PenL26AL30C confirm the higher preference for the formation of CdS 3 O rather than CdS 3 .…”

“…Though we had successfully achieved a fully three-coordinate CdS 3 site, our goal had been to prepare this with Cys. We reconsidered our approach to introducing steric bulk above the Cys plane, and reasoned that by redirecting the Leu side chain towards the Cys site (Figure 10), by use of the D-amino acid, we should exclude water binding [65]. The peptide TRIL12L D L16C (L D ¼ D-Leucine) was prepared and the D-Leu was found to be tolerated and resulted in a well folded coiled coil.…”

2 Methods to Detect and Characterize Metal Ion Binding Sites in RNA

“…The hyperfine interactions of the probe nucleus with the surrounding electric and/or magnetic fields allow to draw conclusions on the closer neighborhood. Originally a method to investigate solids, it has recently been applied to look at metal ion binding in proteins [147,148,[153][154][155][156] ions between two uracil units forming U-Hg(II)-U base pairs in a RNA duplex [157].…”