Metal binding characterization and conformational studies using Raman microscopy of resin-bound poly(aspartic acid).
The metal binding capacities, conditional stability constants, and secondary structure of immobilized polyaspartic acid (PLAsp) (n = 6, 20, and 30) on TentaGel resin were determined when binding Mg2+, Co2+, Cd2+, and Ni2+. Metal binding to the synthesized peptides was evaluated using breakthrough curves from a packed microcolumn and flame atomic absorption spectrophotometry (FAAS) detection. The metal capacities reached values of 590, 2160, and 3710 mumol of metal/g of resin for the 6-mer, 20-mer, and 30-mer, respectively, and this resulted in 2-3 residues per metal for all peptides and metals tested. Surprisingly, the concentrated environment of the resin along with the spatial distribution of attachment groups allowed for most residues to participate in metal binding regardless of the peptide length. Conditional stability constants calculated using single metal binding isotherms indicated that binding strength decreased as the chain length increased on the resin. Raman microscopy on single beads was used to determine PLAsp secondary structure, and all peptides were of a mixed conformation (i.e., beta-sheets, alpha-helices, random chain, etc.) during neutral conditioning and metal binding. Uniquely, the longer 20-mer and 30-mer peptides showed a distinct change from a mixed conformation to beta-sheets and alpha-helices during metal release with acid. This study confirms that metal release by longer immobilized peptides is often assisted by a conformational change, which easily spoils the binding cavity, while shorter peptides may release metal primarily by H+ displacement.
Stair
Jacqueline L
JL
Department of Chemistry and Biochemistry, University of Texas at Austin, Austin, Texas 78712, USA.
Holcombe
James A
JA
D000577:Amides / Q000737:chemistry; D008670:Metals / Q000737:chemistry*; D008853:Microscopy; D010455:Peptides / Q000737:chemistry*; D011487:Protein Conformation; D020539:Sequence Analysis, Protein; D013059:Spectrum Analysis, Raman
D000577:Amides; D008670:Metals; D010455:Peptides; C017645:polyaspartate