Identification of two-histidines one-carboxylate binding motifs in proteins amenable to facial coordination to metals

Abstract

Among natural metalloenzymes, the facial two-histidines one-carboxylate binding motif (FTM) is a widely represented first coordination sphere motif present in the active site of a variety of metalloenzymes. A PDB search revealed a total of 1685 structures bearing such FTMs bound to a metal. Sixty statistically representative FTMs were selected and used as template for the identification of structurally characterized proteins bearing these three amino acids in a propitious environment for binding to a transition metal. This geometrical superposition search, carried out using the STAMPS software, returned 2320 hits. While most consisted of either apo-FTMs or bore strong sequence homology to known FTMs, seven such structures lying within a cavity were identified as novel and viable scaffolds for the creation of artificial metalloenzymes bearing an FTM.

Enzyme repurposing of a hydrolase as an emergent peroxidase upon metal binding

“…For this study, we selected seven scaffolds bearing an HHD/E motif located within a binding pocket predisposed to bind metals. 33 , 34 The search was expanded to include HHN/Q motifs, revealing six additional potential metal binders following a single point mutation. In total, six of the thirteen cloned proteins could be overexpressed in E. coli and purified using a Strep-tag II (Fig.…”

“…Using this approach, we recently reported on the in silico identification of non-metallated two-histidine one-carboxylate metal binding motifs within the structurally characterized proteins of the PDB. 34 Herein we report our efforts to valorise the in silico study by creating an artificial metallo-peroxidase upon addition of a metal cofactor to a protein harboring a latent mononuclear metal binding site, Fig. 1 .…”

Rational design of metalloenzymes: From single to multiple active sites

“…Although more than a half natural proteins are no-metal containing proteins [66], it does not mean that they definitely do not bind metal ions. Ward and co-workers [67] did a systematic search in the protein data bank (PDB) and identified 121 nonmetallated proteins with a potential metal-binding motif of 2-His-1-Glu/Asp. Therefore, these proteins offer idea protein scaffolds for directly incorporating metal ions in the latent metal-binding site, which may confer catalytic activity.…”

Secondary Structure Preferences of Mn2+ Binding Sites in Bacterial Proteins

“…It is known that the “two-histidines-one-carboxylate” binding motif is a widely represented first coordination sphere motif present in the active site of a variety of metalloenzymes [24]. Since histidine and two amino acid residues with carboxyl groups in their side chains are the major binders of Mn 2+ , this motif should be present in our data set as well.…”