Li-June Ming

Immobilization of MP-11 into a Mesoporous Metal–Organic Framework, MP-11@mesoMOF: A New Platform for Enzymatic Catalysis

Microperoxidase-11 has for the first time been successfully immobilized into a mesoporous metal-organic framework (MOF) consisting of nanoscopic cages and it demonstrates superior enzymatic catalysis performances compared to its mesoporous silica counterpart.

How can proteins enter the interior of a MOF? Investigation of cytochrome c translocation into a MOF consisting of mesoporous cages with microporous windows.

It has been demonstrated for the first time that the heme protein cytochrome c (Cyt c) can enter the interior of a MOF despite the larger molecular dimension of the protein relative to the access pore sizes. Mechanistic studies suggest that the Cyt c molecules must undergo a significant conformational change during translocation into the MOF interior through the relatively small nanopores.

Metal binding and structure–activity relationship of the metalloantibiotic peptide bacitracin

Bacitracin is a widely used metallopeptide antibiotic produced by Bacillus subtilis and Bacillus licheniformis with a potent bactericidal activity directed primarily against Gram-positive organisms. This antibiotic requires a divalent metal ion such as Zn(2+) for its biological activity, and has been reported to bind several other transition metal ions, including Mn(2+), Co(2+), Ni(2+), and Cu(2+). Despite the widespread use of bacitracin since its discovery in the early 1940s, the structure-activity relationship of this drug has not been established and the coordination chemistry of its metal complexes was not fully determined until recently. This antibiotic has been suggested to influence cell functioning through more than one route. Since bacterial resistance against bacitracin is still rare despite several decades of widespread use, this antibiotic can serve as an ideal lead for the design of potent peptidyl antibiotics lacking bacterial resistance. In this review, the results of physical (including NMR, EPR, and EXAFS) and molecular biological studies regarding the synthesis and structure of bacitracin, the coordination chemistry of its metal derivatives, the mechanism of its antibiotic actions, its influence on membrane function, and its structure and function relationship are discussed.

Size-Selective Biocatalysis of Myoglobin Immobilized into a Mesoporous Metal–Organic Framework with Hierarchical Pore Sizes

The protein myoglobin has been successfully immobilized into a mesoporous metal-organic framework with hierarchical pore sizes, which demonstrates interesting size-selective biocatalysis as well as superior catalytic activities toward small substrate oxidation compared to its mesoporous silica material counterpart.

THE MECHANISTIC ROLE OF THE COORDINATED TYROSINE IN ASTACIN

Crayfish astacin belongs to the only Zn protein family containing a coordinated Tyr ligand in the active site, and is a rare example of Zn enzymes whose activity can be significantly restored by Cu 2a . The highly active Co 2a and Cu 2a derivatives of astacin can serve as good models of the native enzyme for structural and mechanistic studies by means of optical and magnetic resonance techniques. Upon the introduction of the inhibitor Tyr-hydroxamate to these two metal derivatives of astacin, the coordinated Tyr in the active site is detached based on our optical and NMR studies in solution. The significance of the detachment of the coordinated Tyr in the action of astacin is fourfold: (1) to enhance the Lewis acidity of the active site metal, (2) to balance the negative charge of the transition state gem-diolate-enzyme complex, (3) to relieve the steric crowding upon substrate binding, and (4) to stabilize the enzyme-substrate complex by way of a H-bonding. ” 1998 Published by Elsevier Science Inc. All rights reserved.

Catechol oxidase-like oxidation chemistry of the 1-20 and 1-16 fragments of Alzheimer's disease-related β-amyloid peptide: Their structure-activity correlation and the fate of hydrogen peroxide

The Cu2+ complexes of the 1–16 and the 1–20 fragments of the Alzheimers disease-related β-amyloid peptide (CuAβ) show significant oxidative activities toward a catechol-like substrate trihydroxylbenzene and plasmid DNA cleavage. The latter reflects possible oxidative stress to biological macromolecules, yielding supporting data to the pathological role of these soluble Aβ fragments. The former exhibits enzyme-like kinetics and is dependent on [H2O2], exhibiting kcat of 0.066 s–1 (6000-fold higher than the reaction without CuAβ) and kcat/Km of 37.2 m–1s–1 under saturating [H2O2] of ∼0.24%. This kinetic profile is consistent with metal-centered redox chemistry for the action of CuAβ. A mechanism is proposed by the use of the catalytic cycle of dinuclear catechol oxidase as a working model. Trihydroxylbenzene is also oxidized by CuAβ aerobically without H2O2, affording rate constants of 6.50 × 10–3 s–1 and 3.25 m–1s–1. This activity is also consistent with catechol oxidase action in the absence of H2O2, wherein the substrate binds and reduces the Cu2+ center first, followed by O2 binding to afford the μ-η2:η2-peroxo intermediate, which oxidizes a second substrate to complete the catalytic cycle. A tetragonally distorted octahedral metal coordination sphere with three coordinated His side chains and some specific H-bonding interactions is concluded from the electronic spectrum of CuAβ, hyperfine-shifted 1H NMR spectrum of CoAβ, and molecular mechanics calculations. The results presented here are expected to add further insight into the chemistry of metallo-Aβ, which may assist better understanding of the neuropathology of Alzheimers disease.

Mechanistic studies of the astacin-like Serratia metalloendopeptidase serralysin: highly active (>2000%) Co(II) and Cu(II) derivatives for further corroboration of a "metallotriad" mechanism

Abstract. Serralysin is a bacterial Zn-endopeptidase which has been considered a virulence factor to cause tissue damage and anaphylactic response. It contains a coordinated Tyr that is unique to the astacin-like Zn enzymes. The coordinated Tyr has been proposed to play an important role in the action of this endopeptidase family. Several metal-substituted derivatives of serralysin (including Mn2+, Co2+, Ni2+, Cu2+, and Cd2+ derivatives) are found to exhibit significant activities. Particularly, the Co- and Cu-substituted derivatives exhibit much higher activities than the native serralysin toward the hydrolysis of the tripeptide mimic benzoyl-Arg-p-nitroanilide, i.e., 35 and 49 times higher in kcat and 33 and 26 times in kcat/Km, respectively. Such remarkably higher activities of metal-substituted derivatives, especially the Cu derivative, than that of the native Zn enzyme are rare in the literature, reflecting the uniqueness of this enzyme among all Zn enzymes. The significantly different kcat yet similar Km values among the several metal derivatives suggests that the metal center is involved in catalysis, but not necessarily in the binding of the substrate, whereas the dramatically different inhibition constants for Arg-hydroxamate binding to the metal-substituted derivatives indicates direct binding of this inhibitor to the metal center. The activity-pH profiles of serralysin and its Co2+ and Cu2+ derivatives and the optical-pH profile of Cu-serralysin have been obtained, in which the decrease in activity at higher pH values was found to be associated with a dramatic increase in the Tyr-to-Cu2+ charge transfer transitions. This observation suggests that the binding of Tyr216 to the metal is inhibitory. A metal-centered mechanism is proposed for serralysin catalysis based on the results presented here, in which the detachment of the coordinated Tyr and formation of a H-bond with the transition-state complex are considered essential for the stabilization of the transition state.

Metal-binding and active-site structure of di-zinc Streptomyces griseus aminopeptidase

Abstract Streptomyces griseus aminopeptidase has been characterized to have a dinuclear active site and to follow a dinuclear hydrolytic mechanism by means of activity assay, optical, and NMR spectroscopy. A sequential binding of Co2+ to the dinuclear sites in 20 mM Mes buffer at pH 6.1 has also been established. The results from these studies suggest that the two metal sites have a five-coordination sphere, with at least one coordinated His each. A di-Cu2+-substituted derivative of the enzyme has been prepared which exhibits a 1H NMR spectrum with sharp hyperfine-shifted signals, again indicating the presence of a dinuclear active site. This 1H NMR spectrum with sharp hyperfine-shifted features represents a first of its kind for a di-Cu2+ center in metalloproteins.

A plausible role of salivary copper in antimicrobial activity of histatin-5—Metal binding and oxidative activity of its copper complex

Histatin-5 (Hn5) is an antimicrobial salivary peptide of 24 amino acids. Two specific metal-binding sites were revealed with electronic, NMR, and EPR spectroscopy. The complex Cu(2)(II)-Hn5 effectively oxidizes catechol, exhibiting enzyme-like kinetics (k(cat)=0.011 and 0.060 s(-1) and k(cat)/K(m)=19 and 50 M(-1)s(-1) without and with 12.8mM H(2)O(2), respectively). The significant oxidative activity may contribute to the biological activity of this antibiotic metallopeptide.

1H NMR, mechanism, and mononuclear oxidative activity of the antibiotic metallopeptide bacitracin: The role of d -Glu-4, interaction with pyrophosphate moiety, DNA binding and cleavage, and bioactivity

The peptidyl antibiotic bacitracin (Bc) is one of the most widely used antibiotics which can bind divalent transition metal ions, including Mn(II), Co(II), Ni(II), Cu(II), and Zn(II). The metal binding is essential for its antimicrobial activity. Previous analysis of the hyperfine-shifted (1)H NMR signals of Co(II)-Bc A(1) revealed the structure of the metal binding environment and a potential hydrophobic site important for the bioactivity of this antibiotic. Co(II)-Bc in DMSO shows relatively sharper hyperfine-shifted (1)H NMR signals compared with the spectrum acquired in an aqueous solution, allowing more thorough analysis of the signals with 1D and 2D NMR methods. Pyrophosphate and derivatives bind to Co(II)-Bc to form kinetically inert ternary complexes. The coordinated D-Glu-4 is found detached from the metal center of metallobacitracin upon trimetaphosphate binding, implying its role in the antibiotic activity of Bc. We further demonstrate in this report the structure-function relationship on desamido-Bc of low antibiotic activity by the use of NMR, wherein D-Glu-4 is suggested to be important for the bioactivity of Bc. The interaction of the phospho-moiety with Bc is also reflected by DNA binding, wherein metal-free Bc does not bind DNA, whereas various metal complexes of Bc do. Cu(II)-Bc was further demonstrated to bind and oxidatively cleave DNA under reduction conditions in the air. It also exhibited a significant oxidative activity toward catechol oxidation, showing enzyme-like saturation kinetics with k(cat) = 7.0 x 10(-3) s(-1) and k(cat)/K(m) = 2.1 M(-1) s(-1) aerobically and k(cat) = 0.38 s(-1) and k(cat)/K(m) = 14.7 M(-1) s(-1) in the presence of 32 mM of H(2)O(2). The binding of pyrophosphate moiety to metallobacitracin, the detachment of d-Glu-4, and the significant oxidative activity of Cu(II)-Bc provide further insights into the bioactivity of this metallopeptide and Cu-oxygen chemistry.