Combining Native Mass Spectrometry and Proteomics to Differentiate and Map the Metalloform Landscape in Metallothioneins.

Within the intricate landscape of the proteome, approximately 30% of all proteins bind metal ions. This repertoire is even larger when considering all the different forms of a protein, known as proteoforms. Here, we propose the term "metalloforms" to refer to different structural or functional variations of a protein resulting from the binding of various hetero- or homogeneous metal ions.

Differentiated Zn(II) binding affinities in animal, plant, and bacterial metallothioneins define their zinc buffering capacity at physiological pZn.

Metallothioneins (MTs) are small, Cys-rich proteins present in various but not all organisms, from bacteria to humans. They participate in zinc and copper metabolism, toxic metals detoxification, and protection against reactive species. Structurally, they contain one or multiple domains, capable of binding a variable number of metal ions.

Structural Characterization of Cu(I)/Zn(II)-metallothionein-3 by Ion Mobility Mass Spectrometry and Top-Down Mass Spectrometry.

Mammalian zinc metallothionein-3 (ZnMT3) plays an important role in protecting against copper toxicity by scavenging free Cu(II) ions or removing Cu(II) bound to β-amyloid and α-synuclein. While previous studies reported that ZnMT3 reacts with Cu(II) ions to form Cu(I)Zn(II)MT3ox containing two disulfides (ox), the precise localization of the metal ions and disulfides remained unclear. Here, we undertook comprehensive structural characterization of the metal-protein complexes formed by the reaction between ZnMT3 and Cu(II) ions using native ion mobility mass spectrometry (IM-MS).

OaAEP1 Ligase-Assisted Chemoenzymatic Synthesis of Full Cysteine-Rich Metal-Binding Cyanobacterial Metallothionein SmtA.

Among all approaches used for the semisynthesis of natural or chemically modified products, enzyme-assisted ligation is among the most promising and dynamically developing approaches. Applying an efficient C247A mutant of plant ligase OaAEP1 and solid-phase peptide synthesis chemistry, we present the chemoenzymatic synthesis of a complete sequence of the cysteine-rich and metal-binding cyanobacterial metallothionein Synechococcus metallothionein A (SmtA). Zn(II) and Cd(II) binding to the newly synthesized SmtA showed identical properties to the protein expressed in .