Molar absorption coefficients and stability constants of Zincon metal complexes for determination of metal ions and bioinorganic applications

Kocyła, Anna; Pomorski, Adam; Krężel, Artur

doi:10.1016/j.jinorgbio.2017.08.006

Cited by 44 publications

(43 citation statements)

References 99 publications

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“…A) Cu II binds to the ligand phen with K 1 , β 2 , and β 3 values of 9.0, 15.7, and 20.8, respectively . The complex produces a charge‐transfer band at λ =265 nm ( ϵ ≈ 90 000 m −1 cm −1 ), as previously determined; thus providing a colorimetric handle for measuring competitive Cu II affinities to other species by using similar methods to those described by Kocyła et al . B) Electronic absorption spectra of C‐peptide titrated into solutions of 10 μ m Cu II and 40 μ m phen in 50 m m Bis ⋅ Tris, pH 7.4.…”

Section: Figurementioning

confidence: 80%

“…Titration of C‐peptide into a solution of ZI–Zn II resulted in a decrease of the MLCT band from ZI–Zn II at λ =618 nm, which suggested that C‐peptide effectively competed with ZI for Zn II . Using

K \binom{ZI - Zn^{II}}{d}

, we followed the approach reported by Kocyła et al . and estimated the formation constant of the Zn II /C‐peptide complex to be log K between 4.0 and 5.0 (Figure B and Figure S1 and Table S1 in the Supporting Information).…”

Section: Figurementioning

confidence: 99%

“…A) Zn II binds to the ligand ZI with log K =5.7. The complex produces a metal–ligand charge‐transfer (MLCT) band at λ =618 nm, providing a colorimetric handle for measuring competitive Zn II affinities to other species through methods described by Kocyła et al . B) Electronic absorption spectra of C‐peptide titrated into solutions of 10 μ m Zn II and 20 μ m ZI in 50 m m HEPES, pH 7.4.…”

Section: Figurementioning

confidence: 99%

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Analysis of Metal Effects on C‐Peptide Structure and Internalization

et al. 2019

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The connecting peptide (C‐peptide) has received increased attention for its potential therapeutic effects in ameliorating illnesses such as kidney disease and diabetes. Although the mechanism of C‐peptide signaling remains elusive, evidence supports its internalization and intracellular function. Emerging research is uncovering the diverse biological roles metals play in controlling and affecting the function of bioactive peptides. The work presented herein investigates interactions between C‐peptide and first‐row d‐block transition metals, as well as their effects on C‐peptide internalization into cells. Through spectroscopic techniques, it is demonstrated that CrIII, CuII, and ZnII bind to C‐peptide with differing stoichiometries and biologically relevant affinities. In addition, metal binding elicits both subtle changes in secondary structure and inhibits adoption of an α‐helical character in environments where the dielectric constants are reduced. This study shows how metal ions can modulate peptide hormone activity through subtle structural changes to disrupt cellular uptake.

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Section: Figurementioning

confidence: 80%

K \binom{ZI - Zn^{II}}{d}

Section: Figurementioning

confidence: 99%

Section: Figurementioning

confidence: 99%

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Analysis of Metal Effects on C‐Peptide Structure and Internalization

et al. 2019

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“…When titrated with zinc in buffer alone, 70 μ m Zincon is saturated with 70 μ m ZnCl 2 and an initial increase in absorbance at 620 nm is measured upon addition of 1 μ m ZnCl 2 . Zincon has a K d of 214 n m for a 1 : 1 complex with zinc at pH 8 . However, when competing with 5 μ m apo‐ZnT8 CTD (either variant), the initial increase in absorbance is not seen until 10 μ m Zn 2+ is added, indicating that both ZnT8 CTD variants contain two Zn 2+ ‐binding sites that have a tighter affinity than 214 n m and thus outcompete the zinc binding to Zincon.…”

Section: Resultsmentioning

confidence: 99%

The C‐terminal cytosolic domain of the human zinc transporter ZnT8 and its diabetes risk variant

2018

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A significant aspect of the control of cellular zinc in eukarya is its subcellular re‐distribution. One of the four human vesicular zinc transporters, ZnT8, supplies the millimolar zinc concentrations of insulin granules in pancreatic β‐cells, affecting insulin processing, crystallisation and secretion. ZnT8 has a transmembrane and a C‐terminal cytosolic domain; the latter has important functions and purportedly mediates protein–protein interactions, senses cytosolic zinc and/or channels zinc to the transport site in the transmembrane domain (TMD). A common variant W325R in the C‐terminal domain (CTD) increases the risk to develop type 2 diabetes and affects autoantibody specificity in type 1 diabetes. To investigate the differences between the two protein variants, we purified and biophysically characterised both variants of the ZnT8 CTD [R325 variant of ZnT8 CTD (aa267–369) (ZnT8cR) and W325 variant of ZnT8 CTD (aa267–369) (ZnT8cW)]. The domains fold independently of the TMD. Remarkably, the ZnT8cW variant (diabetes protection in the full‐length protein) is less thermostable than the ZnT8cR variant (diabetes risk in the full‐length protein). The ZnT8cW monomers associate with higher affinity. Both CTD variants bind zinc with a stoichiometry that differs from bacterial homologues, emphasising the limitation of the latter as models for the structure and function of the human proteins. The relatively small but reproducible differences between the two ZnT8 CTD variants begin to provide a molecular basis for the different diabetes susceptibility caused by the full‐length ZnT8 proteins.

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“…Disappearance of the 480 nm peak was used to calculate the percentage of zinc saturation of the Zincon molecule. The dissociation constant of the Zn-Zincon complex is 2.09 · 10 Ϫ6 M, corresponding to a stability constant of log K 1 ϭ 5.89, and no Zn 2 -Zincon complexes were formed (45). At an equimolar zinc-to-Zincon ratio, Zincon was 75% saturated (Fig.…”

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confidence: 99%

Interplay between the Zur Regulon Components and Metal Resistance in Cupriavidus metallidurans

et al. 2019

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The Zur regulon is central to zinc homeostasis in the zinc-resistant bacterium Cupriavidus metallidurans. It comprises the transcription regulator Zur, the zinc importer ZupT, and three members of the COG0523 family of metal-chaperoning G3Etype GTPases, annotated as CobW1, CobW2, and CobW3. The operon structures of the zur and cobW1 loci were determined. To analyze the interplay between the Zur regulon components and metal resistance, deletion mutants were constructed from the wild-type strain CH34 and various other strains. The Zur regulon components interacted with the plasmid-encoded and chromosomally encoded metal resistance factors to acquire metals from complexes of EDTA and for homeostasis of and resistance to zinc, nickel, cobalt, and cadmium. The three G3E-type GTPases were characterized in more detail. CobW1 bound only 1 Zn atom per mol of protein with a stability constant slightly above that of 2-carboxy-2=-hydroxy-5=-sulfoformazylbenzene (Zincon) and an additional 0.5 Zn with low affinity. The CobW1 system was necessary to obtain metals from EDTA complexes. The GTPase CobW2 is a zinc storage compound and bound 0.5 to 1.5 Zn atoms tightly and up to 6 more with lower affinity. The presence of MgGTP unfolded the protein partially. CobW3 had no GTPase activity and equilibrated metal import by ZupT with that of the other metal transport systems. It sequestered 8 Zn atoms per mol with decreasing affinity. The three CobWs bound to the metal-dependent protein FolE IB2 , which is encoded directly downstream of cobW1. This demonstrated an important contribution of the Zur regulon components to metal homeostasis in C. metallidurans. IMPORTANCE Zinc is an important transition metal cation and is present as an essential component in many enzymes, such as RNA polymerase. As with other transition metals, zinc is also toxic at higher concentrations so that living cells have to maintain strict control of their zinc homeostasis. Members of the COG0523 family of metal-chaperoning GE3-type GTPases exist in archaea, bacteria, and eucaryotes, including humans, and they may be involved in delivery of zinc to thousands of different proteins. We used a combination of molecular, physiological, and biochemical methods to demonstrate the important but diverse functions of COG0523 proteins in C. metallidurans, which are produced as part of the Zur-controlled zinc starvation response in this bacterium.

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Molar absorption coefficients and stability constants of Zincon metal complexes for determination of metal ions and bioinorganic applications

Cited by 44 publications

References 99 publications

Analysis of Metal Effects on C‐Peptide Structure and Internalization

Analysis of Metal Effects on C‐Peptide Structure and Internalization

The C‐terminal cytosolic domain of the human zinc transporter ZnT8 and its diabetes risk variant

Interplay between the Zur Regulon Components and Metal Resistance in Cupriavidus metallidurans

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