The small, cysteine-rich metallothionein family of proteins is currently considered to play a critical role in the provision metals metalloenzymes. However, there limited information available on mechanisms these fundamentally important interactions. We report competitive zinc metalation apocarbonic anhydrase presence apometallothionein 1A using electrospray-ionization mass spectrometry. These experiments revealed relative affinities all species solution. carbonic shown compete efficiently...

Metallothioneins (MTs) are cysteine-rich, metal-binding proteins that found throughout Nature. This ubiquity highlights their importance in essential metal regulation, heavy detoxification and cellular redox chemistry. Missing from the current description of MT function is underlying mechanism by which MTs achieve proposed biological functions. To date, there have been conflicting reports on binding structures intermediates formed during metalation apoMTs. The form metal-bound dictates...

Mammalian metallothioneins (MTs) are small, metal binding proteins implicated in cellular ion homeostasis and heavy detoxification. Divalent, metal-saturated MTs form two distinct domains; the N-terminal β domain binds three metals using nine Cys residues, C-terminal α four with 11 residues. Domain selection during zinc cadmium exchange to human MT1A was examined a series of competition reactions mixtures isolated fragments. These experiments were conducted at biologically significant pH...

The flexible coordination stoichiometry of a relatively high number metal ions is property unique to the metallothionein (MT) family proteins. Mammalian MTs, for example, accommodate up seven divalent in tetrahedral geometries, using its complement 20 cysteine ligands. lability metals from these metalloclusters has been used support proposal MTs acting as chaperones, by donating other metal-binding exchange kinetics between human MT1A and carbonic anhydrase (CA) were examined time-dependent...

Copper proteins have the capacity to serve as both redox active catalysts and purely electron transfer centers. A longstanding question in this field is how function of histidine ligated Cu centers are modulated by δ vs ε-nitrogen ligation imidazole. Evaluating impact these coordination modes on structure comparative analysis deposited crystal structures confounded factors such differing protein folds disparate secondary spheres that make direct comparison isomers difficult. Here, we present...

The facile axial ligand exchange properties of gallium(III) protoporphyrin IX in methanol solution were utilized to explore self-association interactions by NMR techniques. Structural changes observed, as well competitive behavior with the ligands acetate and fluoride, which differed from that seen synthetic analogue octaethylporphyrin lacks acid groups its side-chains has less heterogeneity indicated absorption MCD spectroscopies. propionic side chains are implicated all such PPIX, both...

Ga(III)protoporphyrin-IX (Ga-PP) has been proposed as a model for the key interporphyrin interactions in malaria pigment. Unlike paramagnetic parent iron heme derivatives, Ga-PP is readily soluble methanol (MeOH). We report optical, mass spectroscopic, and theoretical results well its reactions with myoglobin. UV–visible absorption MCD spectroscopy show that exhibits typical spectrum main group metal: Q-band at 539 nm B band 406 when dissolved MeOH. also optical data Zn(II)protoporphyrin IX...

Protein design is a powerful tool for interrogating the basic requirements function of metal site in way that allows selective incorporation elements are important function. Rubredoxins small electron transfer proteins with reduction potential centered near 0 mV (vs normal hydrogen electrode). All previous attempts to rubredoxin have focused on incorporating canonical CXXC motifs addition reproducing peptide fold or using flexible loop regions define morphology site. We produced an utterly...

Abstract Bacterial expression of β‐lactamases, which hydrolyze β‐lactam antibiotics, contributes to the growing threat antibacterial drug resistance. Metallo‐β‐lactamases, such as NDM‐1, use catalytic zinc ions in their active sites and nearly all clinically available antibiotics. Inhibitors metallo‐β‐lactamases are urgently needed overcome this resistance mechanism. Zinc‐binding compounds promising leads for inhibitor development, many NDM‐1 inhibitors contain zinc‐binding pharmacophores....

Mammalian metallothioneins (MTs) bind up to seven Zn2+ using a large number of cysteine residues relative their small size and can act as zinc-chaperones. In metal-saturated Zn7–MTs, the zinc ions are co-ordinated tetrahedrally into two distinct clusters separated by linker; N-terminal β-domain [(Zn3Cys9)3−] C-terminal α-domain [(Zn4Cys11)3−]. We report on competitive metalation apo-carbonic anhydrase [CA; metal-free CA (apo-CA)] in presence apo-metallothionein 1A domain fragments identify...

While many life-critical reactions would be infeasibly slow without metal cofactors, a detailed understanding of how protein structure can influence catalytic activity remains elusive. Using de novo designed three-stranded coiled coils (TRI and Grand peptides formed using heptad repeat approach), we examine the insertion three residue discontinuity, known as stammer insert, directly adjacent to (His)3 binding site alters activity. The stammer, which locally twist helix, significantly...

Blue copper proteins have a constrained Cu(II) geometry that has proven difficult to recapitulate outside native cupredoxin folds. Previous work successfully designed green which could be tuned blue using exogenous ligands, but the question of how one can create self-contained site within de novo scaffold, especially removed from fold, remained. We recently reported red protein three helical bundle scaffold we later revisited and determined nitrosocyanin mimic, with CuHis2CysGlu binding...

The pathogenic bacterium Staphylococcus aureus has adopted specialized mechanisms for scavenging iron from its host. nine cell wall and membrane-associated regulated surface determinant (Isd) proteins (IsdH, IsdB, IsdA, IsdC, IsdDEF, IsdG IsdI) allow to scavenge the heme in hemoglobin haptoglobin–hemoglobin. Of these, it is IsdE that chaperones ATP binding cassette-type transmembrane transporter (IsdF). IsdH, IsdA IsdC contain at least one Near Transporter (NEAT) domain. Previous studies...

Abstract Die Frage nach der Beziehung zwischen Struktur und Funktion von Proteinen ist eines größten Rätsel Biochemie. Das De‐novo‐Design Metalloproteinen bietet die Möglichkeit, neu zu bestimmen, was nötig ist, um Grund auf Funktionalität in einer aufzubauen, nicht des natürlichen Vorbilds abgeleitet ist. Dieser Aufsatz konzentriert sich Arbeiten zum Proteindesign, De‐novo‐Metalloproteine innerhalb alpha‐helikaler Gerüststrukturen liefern. Beispiele umfassen De‐novo‐Proteine mit...

Long interspersed nuclear elements-1 (L1) are autonomous retrotransposons that encode two proteins in different open reading frames (ORF1 and ORF2). The ORF1p, which may be an RNA binding chaperone protein, contains a three-stranded coiled coil (3SCC) domain facilitates the formation of biologically active homotrimer. This 3SCC is composed seven amino acid (heptad) repeats as found native designed peptides stammer modifies helical structure. Cysteine residues occur at three hydrophobic...

The human long interspersed nuclear element 1 (LINE1) has been implicated in numerous diseases and suggested to play a significant role genetic evolution. Open reading frame protein (ORF1p) is one of the two proteins encoded this self-replicating mobile element, both which are essential for retrotransposition. structure three-stranded coiled-coil domain ORF1p was recently solved showed presence tris-cysteine layers interior that could function as metal binding sites. Here, we demonstrate...

The rational design and functionalization of small, simple, stable peptides scaffolds is an attractive avenue to mimic catalytic metal-centres complex proteins, relevant for the metalloenzymes with environmental, biotechnological health impacts. de novo designed α

Abstract While many life‐critical reactions would be infeasibly slow without metal cofactors, a detailed understanding of how protein structure can influence catalytic activity remains elusive. Using de novo designed three‐stranded coiled coils ( TRI and Grand peptides formed using heptad repeat approach), we examine the insertion three residue discontinuity, known as stammer insert, directly adjacent to (His) 3 binding site alters activity. The stammer, which locally twist helix,...