scholarly journals Zinc Ion-induced Domain Organization in Metallo-β-lactamases

2009 ◽  
Vol 284 (24) ◽  
pp. 16419-16431 ◽  
Author(s):  
Nathalie Selevsek ◽  
Sandrine Rival ◽  
Andreas Tholey ◽  
Elmar Heinzle ◽  
Uwe Heinz ◽  
...  
Keyword(s):  
Metal Binding ◽  
Metal Ion ◽  
Protein Complexes ◽  
Consensus Sequence ◽  
Zinc Ion ◽  
Ionization Mass ◽  
Protein Ratio ◽  
Metal Free ◽  
Intrinsically Disordered ◽  
Flight Mass Spectrometry

The reversible unfolding of metallo-β-lactamase from Chryseobacterium meningosepticum (BlaB) by guanidinium hydrochloride is best described by a three-state model including folded, intermediate, and unfolded states. The transformation of the folded apoenzyme into the intermediate state requires only very low denaturant concentrations, in contrast to the Zn2-enzyme. Similarly, circular dichroism spectra of both BlaB and metallo-β-lactamase from Bacillus cereus 569/H/9 (BcII) display distinct differences between metal-free and Zn2-enzymes, indicating that the zinc ions affect the folding of the proteins, giving a larger α-helix content. To identify the regions of the protein involved in this zinc ion-induced change, a hydrogen deuterium exchange study with matrix-assisted laser desorption ionization tandem time of flight mass spectrometry on metal-free and Zn1- and Zn2-BcII was carried out. The region spanning the metal binding metallo-β-lactamases (MBL) superfamily consensus sequence His-X-His-X-Asp motif and the loop connecting the N- and C-terminal domains of the protein undergoes a zinc ion-dependent structural change between intrinsically disordered and ordered states. The inherent flexibility even appears to allow for the formation of metal ion-bridged protein-protein complexes which may account for both electrospray ionization-mass spectroscopy results obtained upon variation of the zinc/protein ratio and stoichiometry-dependent variations of 199mHg-perturbed angular correlation of γ-rays spectroscopic data. We suggest that this flexible “zinc arm” motif, present in all the MBL subclasses, is disordered in metal-free MBLs and may be involved in metal ion acquisition from zinc-carrying molecules different from MBL in an “activation on demand” regulation of enzyme activity.

scholarly journals Spectroscopic Studies of Copper and Silver Binding to Metallothioneins

1999 ◽  
Vol 6 (4-5) ◽  
pp. 277-290 ◽  
Author(s):  
Martin J. Stillman
Keyword(s):  
Spectral Data ◽  
Metal Binding ◽  
Emission Spectra ◽  
Structural Data ◽  
Spectroscopic Studies ◽  
Yeast Cells ◽  
Luminescence Spectra ◽  
Ionization Mass ◽  
Mass Spectral ◽  
Metal Free

Mammalian metallothionein is remarkable in its metal binding properties: well-characterized species exist for metal to sulfur ratios of M7S20, M12S20, and M18S20, where M = Cd(ll), Zn(ll), Hg(ll), Ag(I), Au(I), and Cu(I). Circular dichroism and luminescence spectra provide rich details of a complicated metal binding chemistry when metals are added directly to the metal free- or zinc-containing protein. CD spectral data unambiguously identify key metal to protein stoichiometric ratios that result in well-defined structures. Emission spectra in the 450-750 nm region have been reported for metallothioneins containing Ag(I), Au(I), and Cu(I). The luminescence of Cu-MT can also be detected directly from mammalian and yeast cells. Qualitative and quantitative interpretations show that the final structure adopted by Ag-MT is not the same as that formed by Cu(I) ions in Cu-MT. XAFS structural data are reported for a number of metallothioneins, including Ag12-MT and Ag17-MT. Electrospray ionization mass spectrometry provides details on the species formed when Ag(I) binds to metallothionein. Mass spectral data are reported for metal-free MT 2A and Agn-MT (n = 14-18).

Novel 4,4′-{(diphenylmethylene) bis(4,1-phenylene)bis(oxy)}-bridged ball-type phthalocyanines: Electrochemical, electrocatalytic oxygen reducing and heavy metals ions sensing properties

2016 ◽  
Vol 20 (08n11) ◽  
pp. 1319-1333 ◽  
Author(s):  
Elvan Doğan ◽  
Metin Özer ◽  
Ahmet Altındal ◽  
Ali Rıza Özkaya ◽  
Bekir Salih ◽  
...  
Keyword(s):  
Heavy Metal ◽  
Metal Ions ◽  
Quartz Crystal Microbalance ◽  
Metal Ion ◽  
Heavy Metal Ions ◽  
Quartz Crystal ◽  
Structural Difference ◽  
Ionization Mass ◽  
Metal Free ◽  
Quartz Crystal Microbalance Sensor

Novel 4,4′-{(diphenylmethylene)bis(4,1-phenylene)bis(oxy)} bridged ball-type metal-free, zinc(II), cobalt(II) and iron(II) ball-type metallophthalocyanines were achieved by the reaction of the bisphthalonitrile derivative in 2-dimetylaminoethanol. The isolation of the metal-free and metallophthalocyanines were carried out by both planar and column chromatography and also by hot soxhalet extraction. Their structures have been characterized by infrared, ultraviolet-visible,1H nuclear magnetic resonance and matrix assisted laser desorption/ionization mass spectroscopies. The redox characters of the ball-type metallophthalocyanines have been investigated by cyclic voltammetry, square wave voltammetry, controlled potential coulometry and spectroelectrochemistry in nonaqueous media. The investigation suggested that the ball-type complexes form ring-based and/or metal-based mixed-valence species as a result of the remarkable interaction between the two Pc rings and/or metal centers. Furthermore, these species were found to be stable as evidenced by the splitting of the relevant redox processes. The electrocatalytic oxygen reducing performances of the dinuclear ball-type complexes were also studied. The compound involving Fe(II) centers at the phthalocyanine cores displayed higher catalytic performance towards oxygen reduction than those of other ones. By using these compounds as sensing materials, a flow type quartz crystal microbalance sensor was developed for the detection of small concentrations of heavy metal ions. A cadmium ion sensitivity of 2.85 × 104Hz/mg.L[Formula: see text] was observed with ball-type iron(II) phthalocyanine coated quartz crystal microbalance sensor. Detection of other metal ions including Fe[Formula: see text], Zn[Formula: see text], Ag[Formula: see text] and Sn[Formula: see text] were also performed. The results indicated that 1– 4 functionalized quartz crystal microbalance sensors can be used for the detection of heavy metal ions in aqueous solution. Partition coefficients obtained from the linear response regimes of the calibration curves are in the 8.6 × 104–5.4 × 105range. Results show that the minor structural difference between metal free and metallophthalocyanine poses significant impact on metal ion partitioning.

scholarly journals Mutational analysis of the zinc- and substrate-binding sites in the CphA metallo-β-lactamase from Aeromonas hydrophila

2008 ◽  
Vol 414 (1) ◽  
pp. 151-159 ◽  
Author(s):  
Carine Bebrone ◽  
Christine Anne ◽  
Frédéric Kerff ◽  
Gianpiero Garau ◽  
Kris De Vriendt ◽  
...  
Keyword(s):  
Significant Role ◽  
Metal Binding ◽  
Aeromonas Hydrophila ◽  
Metal Ion ◽  
Mutational Analysis ◽  
Substrate Binding ◽  
Zinc Ion ◽  
Negative Effect ◽  
Activity Spectrum ◽  
Substrate Binding Sites

The subclass B2 CphA (Carbapenemase hydrolysing Aeromonas) β-lactamase from Aeromonas hydrophila is a Zn2+-containing enzyme that specifically hydrolyses carbapenems. In an effort to evaluate residues potentially involved in metal binding and/or catalysis (His118, Asp120, His196 and His263) and in substrate specificity (Val67, Thr157, Lys224 and Lys226), site-directed mutants of CphA were generated and characterized. Our results confirm that the first zinc ion is in interaction with Asp120 and His263, and thus is located in the ‘cysteine’ zinc-binding site. His118 and His196 residues seem to be interacting with the second zinc ion, as their replacement by alanine residues has a negative effect on the affinity for this second metal ion. Val67 plays a significant role in the binding of biapenem and benzylpenicillin. The properties of a mutant with a five residue (LFKHV) insertion just after Val67 also reveals the importance of this region for substrate binding. This latter mutant has a higher affinity for the second zinc ion than wild-type CphA. The T157A mutant exhibits a significantly modified activity spectrum. Analysis of the K224Q and N116H/N220G/K224Q mutants suggests a significant role for Lys224 in the binding of substrate. Lys226 is not essential for the binding and hydrolysis of substrates. Thus the present paper helps to elucidate the position of the second zinc ion, which was controversial, and to identify residues important for substrate binding.

Structures of c-di-GMP/cGAMP degrading phosphodiesterase VcEAL: identification of a novel conformational switch and its implication

2019 ◽  
Vol 476 (21) ◽  
pp. 3333-3353 ◽  
Author(s):  
Malti Yadav ◽  
Kamalendu Pal ◽  
Udayaditya Sen
Keyword(s):  
Active Site ◽  
Metal Binding ◽  
Metal Ion ◽  
Triosephosphate Isomerase ◽  
Catalytic Mechanism ◽  
Degradation Mechanism ◽  
Structural Basis ◽  
Conserved Residues ◽  
Phosphodiester Bond ◽  
Cyclic Dinucleotides

Cyclic dinucleotides (CDNs) have emerged as the central molecules that aid bacteria to adapt and thrive in changing environmental conditions. Therefore, tight regulation of intracellular CDN concentration by counteracting the action of dinucleotide cyclases and phosphodiesterases (PDEs) is critical. Here, we demonstrate that a putative stand-alone EAL domain PDE from Vibrio cholerae (VcEAL) is capable to degrade both the second messenger c-di-GMP and hybrid 3′3′-cyclic GMP–AMP (cGAMP). To unveil their degradation mechanism, we have determined high-resolution crystal structures of VcEAL with Ca2+, c-di-GMP-Ca2+, 5′-pGpG-Ca2+ and cGAMP-Ca2+, the latter provides the first structural basis of cGAMP hydrolysis. Structural studies reveal a typical triosephosphate isomerase barrel-fold with substrate c-di-GMP/cGAMP bound in an extended conformation. Highly conserved residues specifically bind the guanine base of c-di-GMP/cGAMP in the G2 site while the semi-conserved nature of residues at the G1 site could act as a specificity determinant. Two metal ions, co-ordinated with six stubbornly conserved residues and two non-bridging scissile phosphate oxygens of c-di-GMP/cGAMP, activate a water molecule for an in-line attack on the phosphodiester bond, supporting two-metal ion-based catalytic mechanism. PDE activity and biofilm assays of several prudently designed mutants collectively demonstrate that VcEAL active site is charge and size optimized. Intriguingly, in VcEAL-5′-pGpG-Ca2+ structure, β5–α5 loop adopts a novel conformation that along with conserved E131 creates a new metal-binding site. This novel conformation along with several subtle changes in the active site designate VcEAL-5′-pGpG-Ca2+ structure quite different from other 5′-pGpG bound structures reported earlier.

scholarly journals Ion Imaging of Native Protein Complexes Using Orthogonal Time-of-Flight Mass Spectrometry and a Timepix Detector

2021 ◽  
Vol 32 (2) ◽  
pp. 569-580
Author(s):  
Anjusha Mathew ◽  
Ronald Buijs ◽  
Gert B. Eijkel ◽  
Frans Giskes ◽  
Andrey Dyachenko ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Protein Complexes ◽  
Time Of Flight ◽  
Ion Imaging ◽  
Native Protein ◽  
Flight Mass Spectrometry

scholarly journals Acylated Anthocyanins from Red Cabbage and Purple Sweet Potato Can Bind Metal Ions and Produce Stable Blue Colors

2021 ◽  
Vol 22 (9) ◽  
pp. 4551
Author(s):  
Julie-Anne Fenger ◽  
Gregory T. Sigurdson ◽  
Rebecca J. Robbins ◽  
Thomas M. Collins ◽  
M. Mónica Giusti ◽  
...  
Keyword(s):  
Metal Ions ◽  
Sweet Potato ◽  
Metal Binding ◽  
Metal Ion ◽  
High Resolution Mass Spectrometry ◽  
Large Set ◽  
Water Addition ◽  
Red Cabbage ◽  
Aluminum Ions ◽  
Purple Sweet Potato

Red cabbage (RC) and purple sweet potato (PSP) are naturally rich in acylated cyanidin glycosides that can bind metal ions and develop intramolecular π-stacking interactions between the cyanidin chromophore and the phenolic acyl residues. In this work, a large set of RC and PSP anthocyanins was investigated for its coloring properties in the presence of iron and aluminum ions. Although relatively modest, the structural differences between RC and PSP anthocyanins, i.e., the acylation site at the external glucose of the sophorosyl moiety (C2-OH for RC vs. C6-OH for PSP) and the presence of coordinating acyl groups (caffeoyl) in PSP anthocyanins only, made a large difference in the color expressed by their metal complexes. For instance, the Al3+-induced bathochromic shifts for RC anthocyanins reached ca. 50 nm at pH 6 and pH 7, vs. at best ca. 20 nm for PSP anthocyanins. With Fe2+ (quickly oxidized to Fe3+ in the complexes), the bathochromic shifts for RC anthocyanins were higher, i.e., up to ca. 90 nm at pH 7 and 110 nm at pH 5.7. A kinetic analysis at different metal/ligand molar ratios combined with an investigation by high-resolution mass spectrometry suggested the formation of metal–anthocyanin complexes of 1:1, 1:2, and 1:3 stoichiometries. Contrary to predictions based on steric hindrance, acylation by noncoordinating acyl residues favored metal binding and resulted in complexes having much higher molar absorption coefficients. Moreover, the competition between metal binding and water addition to the free ligands (leading to colorless forms) was less severe, although very dependent on the acylation site(s). Overall, anthocyanins from purple sweet potato, and even more from red cabbage, have a strong potential for development as food colorants expressing red to blue hues depending on pH and metal ion.

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