Alternative zinc binding peptides as potential tags for recombinant protein purification

Efficient binding of peptides and proteins to metal-chelating resins is a cornerstone of modern biochemical purification. This study evaluates a novel heptapeptide sequence, acetyl-Aa₁-Aa₂-Gly₃-Pro₄-Aa₅-His₆-Cys₇, where Aa₁ = His₁ or Asp₁, Aa₂ = Cys₂ or Asp₂ and Aa₅ = Tyr₅ or Gly₅ for its capacity to bind zinc-chelating resin consisting of divalent zinc chelated by iminodiacetate coupled to 6 % cross-linked agarose beads. Comparisons were made against the widely utilized 7 × His tag using an internal standard method with ion mobility – mass spectrometry analyses and ultra-violet absorption analyses, which quantified the binding efficiency and selectivity of these peptides under pH 8 conditions and the elution from the zinc resin using pH 3.9 or excess imidazole. Results revealed that the heptapeptides acetyl-His₁-Cys₂-Gly₃-Pro₄-Tyr₅-His₆-Cys₇, exhibited binding performance to the zinc chelating resin with conditions commonly used with immobilized metal affinity chromatography and efficient elution from the zinc resin at pH 3.9 or with excess imidazole, suggesting that this heptapeptide has potential as an alternative to polyhistidine tags in affinity-based purification workflows.