The distribution in native populations from Mexico and Central America of the C677T variant in the MTHFR gene.

OBJECTIVES: To explore evolutionary hypotheses for the high frequencies of a substitution in the methylenetetrahydrofolate reductase (MTHFR) gene, in Mexican and Central American Indigenous populations. MATERIALS AND METHODS: We obtained allele frequencies for the C677T variant in the MTHFR gene and ecological information for 37 indigenous samples from Mexico and Central America. We calculated Hardy-Weinberg equilibrium and computed Fst statistics. We computed correlations between the samples' allele frequencies and ecological and geochemical variables. RESULTS: Many of the samples have extremely high frequencies of the T allele ( q ¯  = 0.62, median = 0.66). In this region, the frequency of the T allele decreases from Southeast to Northwest and is significantly correlated with longitude, latitude, altitude, and insolation. CONCLUSIONS: The native people of Central America and Mexico evolved high frequencies of an allele which has been shown to produce deleterious clinical effects including neural tube effects, cardiovascular events, and cancer. This allele has a clinal distribution in the region, perhaps associated with solar irradiation. As (Contreras-Cubas et al., 2016) noted, the traditional diet of these populations, which is high in folate, has likely mitigated the negative effect of the allele. It is of primary importance that their rights to their homeland and traditional diets be respected. It is a matter of Public Health to investigate whether this allele is a factor in the current wave of cardiovascular diseases affecting the majority population of this region, since it descends from the Native peoples and the Mediterranean population, which also has high frequencies of the allele.

Introducing Seven Transition Metal Ions into Terpyridine-Based Supramolecules: Self-Assembly and Dynamic Ligand Exchange Study.

In coordination-driven self-assembly, 2,2':6',2″-terpyridine (tpy) has gained extensive attention in constructing supramolecular architectures on the basis of ⟨tpy-M-tpy⟩ connectivity. In direct self-assembly of large discrete structures, however, the metal ions were mainly limited to Cd(II), Zn(II), and Fe(II) ions. Herein, we significantly broaden the spectrum of metal ions with seven divalent transition metal ions M(II) (M = Mn, Fe, Co, Ni, Cu, Zn, Cd) to assemble a series of supramolecular fractals. In particular, Mn(II), Co(II), Ni(II), and Cu(II) were reported for the first time to form such large and discrete structures with ⟨tpy-M-tpy⟩ connectivity. In addition, the structural stabilities of those supramolecules in the gas phase and the kinetics of the ligand exchange process in solution were investigated using mass spectrometry. Such a fundamental study gave the relative order of structural stability in the gas phase and revealed the inertness of coordination in solution depending on the metal ions. Those results would guide the future study in tpy-based supramolecular chemistry in terms of self-assembly, characterization, property, and application.

Right-Handed Helical Foldamers Consisting of De Novo d-AApeptides.

New types of foldamer scaffolds are formidably challenging to design and synthesize, yet highly desirable as structural mimics of peptides/proteins with a wide repertoire of functions. In particular, the development of peptidomimetic helical foldamers holds promise for new biomaterials, catalysts, and drug molecules. Unnatural l-sulfono-γ-AApeptides were recently developed and shown to have potential applications in both biomedical and material sciences. However, d-sulfono-γ-AApeptides, the enantiomers of l-sulfono-γ-AApeptides, have never been studied due to the lack of high-resolution three-dimensional structures to guide structure-based design. Herein, we report the first synthesis and X-ray crystal structures of a series of 2:1 l-amino acid/d-sulfono-γ-AApeptide hybrid foldamers, and elucidate their folded conformation at the atomic level. Single-crystal X-ray crystallography indicates that this class of oligomers folds into well-defined right-handed helices with unique helical parameters. The helical structures were consistent with data obtained from solution 2D NMR, CD studies, and molecular dynamics simulations. Our findings are expected to inspire the structure-based design of this type of unique folding biopolymers for biomaterials and biomedical applications.

Insights into SOD1-linked amyotrophic lateral sclerosis from NMR studies of Ni(2+)- and other metal-ion-substituted wild-type copper-zinc superoxide dismutases.

The dimeric Cu-Zn superoxide dismutase (SOD1) is a particularly interesting system for biological inorganic chemical studies because substitutions of the native Cu and/or Zn ions by a nonnative metal ion cause minimal structural changes and result in high enzymatic activity for those derivatives with Cu remaining in the Cu site. The pioneering NMR studies of the magnetically coupled derivative Cu2Co2SOD1 by Ivano Bertini and coworkers are of particular importance in this regard. In addition to Co(2+), Ni(2+) is a versatile metal ion for substitution into SOD1, showing very little disturbance of the structure in Cu2Ni2SOD1 and acting as a very good mimic of the native Cu ion in Ni2Zn2SOD1. The NMR studies presented here were inspired by and are indebted to Ivano Bertini's paramagnetic NMR pursuits of metalloproteins. We report Ni(2+) binding to apo wild-type SOD1 and a time-dependent Ni(2+) migration from the Zn site to the Cu site, and the preparation and characterization of Ni2Ni2SOD1, which shows coordination properties similar to those of Cu2Cu2SOD1, namely, an anion-binding property different from that of the wild type and a possibly broken bridging His. Mutations in the human SOD1 gene can cause familial amyotrophic lateral sclerosis (ALS), and mutant SOD1 proteins with significantly altered metal-binding behaviors are implicated in causing the disease. We conclude by discussing the effects of the ALS mutations on the remarkable stabilities and metal-binding properties of wild-type SOD1 proteins and the implications concerning the causes of SOD1-linked ALS.

Metal binding of flavonoids and their distinct inhibition mechanisms toward the oxidation activity of Cu2+-ß-amyloid: not just serving as suicide antioxidants!

The accumulation of plagues of ß-amyloid (Aß) peptides in the brain is a hallmark of Alzheimer's disease (AD). The redox-active Cu and Fe complexes of Aß can cause damage to the neurons potentially via reactive oxygen species (ROS). The significant metal-mediated oxidative activity of CuAß suggests that its presence can be chemically devastating regardless whether it is a cause or a result of AD. Flavonoids exhibit various benefits to human health, attributable to their metal-binding and antioxidation activities to certain extents. Despite broad interests and extensive studies of their metal-binding properties and anti/pro-oxidation activities, these properties and the mechanisms of the activities toward metal-centered oxidation reactions have not been fully revealed and concluded. We report herein distinctive antioxidation mechanisms between two flavonoid families toward the oxidation reactions by CuAß(1-20), wherein the flavonols quercetin (Qr) and myricetin (Mr) competitively inhibit the oxidation of catechol by CuAß(1-20) with K(i) of 11.2 and 32.6 µM, respectively, whereas the flavanols catechin (Ct) and epicatechin (Et) are substrates with k(cat) = 1.01 × 10(-2) and 1.55 × 10(-3) s(-1) and K(m) = 0.94 and 0.55 mM, respectively. Qr has a nearly 10-fold higher antioxidative efficacy than Ct against the oxidation activity of CuAß, while Ct is effectively oxidized, which further decreases its antioxidant capacity. Similar inhibition patterns are observed toward oxidation of the catecholamine neurotransmitter dopamine by CuAß(1-20). Metal ions and CuAß bind Qr with a 1:1 ratio under our experimental conditions through the α-ketoenolate moiety as determined by the use of Co(2+) and Yb(3+) as paramagnetic NMR probes. Unlike flavanols, which are merely suicide antioxidative substrates, flavonols bind to the metal center and prevent metal-mediated redox reactions. We suggest flavonols may serve as leads for drug discovery and/or as agents toward preventing metal-mediated oxidative stress due to AD and other disorders. Moreover, CuAß shows 8.6- and 4.2-fold higher kinetic regioselectivity in terms of k(cat) and k(cat)/K(m), respectively, toward the peroxidation of Ct than that of the enantiomer Et, suggesting potential development of metallo-catalysts in regioselective catalysis by the use of metallopeptides as templates.

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.

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.

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.

Vitamin B6s inhibit oxidative stress caused by Alzheimer's disease-related Cu(II)-ß-amyloid complexes-cooperative action of phospho-moiety.

Cu(II) complexes of Alzheimer's disease-related ß-amyloid (Aß) peptides exhibit metal-centered oxidation chemistry. The metallo-Aß complexes are the hallmark of the disease and have been attributed to the generation of reactive oxygen species (ROS), causing oxidative stress. In this communication, the inhibitions of the oxidative activity of Cu(II)-Aß by vitamin B6 compounds pyridoxamine (PM), pyridoxine (PN), pyridoxal (PL), and pyridoxal-5'-phosphate (PLP) are presented. These B6's are competitive inhibitors toward dopamine oxidation by Cu(II)-Aß(1-20), with K(i) values of 1.4, 8.3, 1.2, and 0.2mM, respectively. The phospho-moiety in PLP seems to exhibit cooperative inhibition, affording a clue for future design of inhibitors.

The folding propensity of α/sulfono-γ-AA peptidic foldamers with both left- and right-handedness.

The discovery and application of new types of helical peptidic foldamers have been an attractive endeavor to enable the development of new materials, catalysts and biological molecules. To maximize their application potential through structure-based design, it is imperative to control their helical handedness based on their molecular scaffold. Herein we first demonstrate the generalizability of the solid-state right-handed helical propensity of the 413-helix of L-α/L-sulfono-γ-AA peptides that as short as 11-mer, using the high-resolution X-ray single crystallography. The atomic level folding conformation of the foldamers was also elucidated by 2D NMR and circular dichroism under various conditions. Subsequently, we show that the helical handedness of this class of foldamer is controlled by the chirality of their chiral side chains, as demonstrated by the left-handed 413-helix comprising 1:1 D-α/D-sulfono-γ-AA peptide. In addition, a heterochiral coiled-coil-like structure was also revealed for the first time, unambiguously supporting the impact of chirality on their helical handedness. Our findings enable the structure-based design of unique folding biopolymers and materials with the exclusive handedness or the racemic form of the foldamers in the future.

Methionine does not reduce Cu(II)-beta-amyloid!--rectification of the roles of methionine-35 and reducing agents in metal-centered oxidation chemistry of Cu(II)-beta-amyloid.

The potential risk of metal-centered oxidative catalysis has been overlooked in the research of the copper complexes of the Alzheimer's disease-related beta-amyloid (Abeta) peptides. Cu(2+) complexes of Abeta(1-40) and its 1-16 and 1-20 fragments have recently been shown to exhibit significant metal-centered oxidative activities toward several catecholamine neurotransmitters with and without H(2)O(2) around neutral pH [G.F.Z. da Silva, L.-J. Ming, "Metallo-ROS" in Alzheimer's disease: metal-centered oxidation of neurotransmitters by Cu(II)-beta-amyloid and neuropathology of Alzheimer's disease, Angew. Chem. Int. Ed. 46 (2007) 3337-3341]. The results further support the metallo-Abeta-associated oxidative stress theory often considered to be connected to the neuropathology of the disease. The metal-centered oxidative catalysis of CuAbeta(1-16/20) challenges the long-standing proposed redox role of Met35 in Abeta because Abeta(1-16/20) do not contain a Met. External Met has been determined by kinetic, optical, and electron paramagnetic resonance methods to bind directly to the Cu(2+) center of CuAbeta(1-40) and CuAbeta(1-20) with K(d)=2.8 mM and 11.3 microM, respectively, which reflects less accessibility of the metal center in the full-length CuAbeta(1-40). However, Met does not serve as a reducing agent for the Cu(II) which thus must amplify the observed oxidative catalysis of CuAbeta(1-20)through a non-redox mechanism. Conversely, the CuAbeta-catalyzed oxidation reaction of dopamine is inhibited by bio-available reducing agents such as ascorbate (competitive K(ic)=66 microM) and glutathione (non-competitive, K(inc)=53 microM). These data indicate that the oxidation chemistry of metallo-Abeta is not initiated by Met35. The results yield further molecular and mechanistic insights into the roles of metallo-Abeta in this disease.

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.

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.

Purification and characterization of extracellular lipases from Pseudomonas monteilii TKU009 by the use of soybeans as the substrate.

A lipase-producing bacterium was isolated and identified as Pseudomonas monteilii TKU009. A lipase (F2) and lipase-like materials (F1) were purified from the culture supernatant of P. monteilii TKU009 with soybean powder as the sole carbon/nitrogen source. The molecular mass of F1 and F2 was estimated to be 44 kDa by SDS-PAGE and gel filtration. The optimum pH, optimum temperature, and pH and thermal stabilities of F2 were 7, 40 degrees C, 8-11, and 50 degrees C; and of F1 were 6, 40 degrees C, 6-7, and 50 degrees C, respectively. F2 was completely inhibited by EDTA and slightly by Mg(2+), Fe(2+), Mn(2+), and SDS. F1 was completely inhibited by EDTA and Fe(2+) and strongly by Zn(2+), Mn(2+), Ca(2+), Mg(2+), and SDS. The activities of both the enzymes were enhanced by the addition of non-ionic surfactants Triton X-100 and Tween 40, especially for F1. F2 preferably acted on substrates with a long chain (C10-C18) of fatty acids, while F1 showed a broad spectrum on those with chain length of C4-C18. The marked activity of F2 in organic solvents makes it an ideal choice for application in a water-restricted medium including organic synthesis.

Comprehensive 2D (1)H NMR Studies of Paramagnetic Lanthanide(III) Complexes of Anthracycline Antitumor Antibiotics.

The binding of several lanthanide(III) ions to anthracycline antitumor antibiotics daunomycin and adriamycin in methanol and aqueous solutions has been studied by means of optical and 2D NMR (COSY, TOCSY, and EXSY) techniques. These results indicate that a 1:1 Yb(3+)-drug complex (1) is the predominant complex at a metal-to-ligand ratio <10 with slightly higher proton activities, e.g., approximately pH 4-5 in an aqueous solution. In the presence of a base, a 1:2 (2) or 1:3 (3) Yb(3+)-drug complex can be formed. In addition, a 2:1 complex (4) is formed when the metal-to-drug ratio is >25. These Yb(3+)-drug complexes undergo slow chemical exchange with each other relative to the NMR time scale. Therefore, 1D and 2D magnetization transfer experiments can be utilized for the assignment of the isotropically shifted signals arising from the drug nuclei in the various paramagnetic complexes. The spin-lattice (T(1)) relaxation times and solution magnetic susceptibilities of these Yb(3+)-drug complexes confirmed the binding of the metal ion to 11,12-beta-ketophenolate in all the complexes (except the second Yb(3+) in the 2:1 complex which binds to the 5,6-beta-ketophenolate). Several other lanthanide(III) ions Pr(3+), Eu(3+), and Dy(3+) show similar binding properties to daunomycin based on optical and NMR studies. The binding of Yb(3+) to daunomycin has a profound effect on the reduction potential of the drug, showing a decrease in the potential by 150 mV upon addition of 1 equiv of Yb(3+) to the drug solution. This observation indicates that metal ions must play a significant role in the action of these family of drugs in vivo.

NMR Study of Dendrimer Structures Using Paramagnetic Cobalt(II) as a Probe.

Cobalt(II) has been utilized as an external paramagnetic (1)H NMR probe for the study of the structure of dendrimers that possess specifically located metal recognition sites. The isotropically shifted (1)H NMR signals of the Co(II) complexes of two 2,6-diamidopyridine-containing dendrimers have been fully assigned by means of 1D and 2D NMR techniques, including NOE difference, EXSY, COSY, and TOCSY. T(1) values of the isotropically shifted signals were used to calculate metal-proton distances to build a molecular model of the internal structure of the dendrimers. The presence of sizable cavities within the dendrimers was observed, including a loosely packed conformation for the 2,6-diamidopyridine moiety to bind to potential guest molecules.

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.

Mechanistic role of each metal ion in Streptomyces dinuclear aminopeptidase: PEPTIDE hydrolysis and 7x10(10)-fold rate enhancement of phosphodiester hydrolysis.

The dinuclear aminopeptidase from Streptomyces griseus (SgAP) and its metal derivatives catalyze the hydrolysis of the phosphoester bis(p-nitrophenyl) phosphate (BNPP) and the phosphonate ester p-nitrophenyl phenylphosphonate with extraordinary rate enhancements at pH 7.0 and 25 degrees C [A. Ercan, H. I. Park, L.-J. Ming, Biochemistry 45, (2006) 13779-13793.], reaching 6.7 billion-fold in terms of the first-order rate constant of the di-Co(II) derivative with respect to the autohydrolytic rates. Since phosphoesters are transition state-like inhibitors in peptide hydrolysis, their hydrolysis by SgAP is quite novel. Herein, we report the investigation of this proficient alternative catalysis of SgAP and the role of each metal ion in the dinuclear site toward peptide and BNPP hydrolysis. Mn(II) selectively binds to one of the dinuclear metal sites (M1), affording MnE-SgAP with an empty (E) second site for the binding of another metal (M2), including Mn(II), Co(II), Ni(II), Zn(II), and Cd(II). Peptide hydrolysis is controlled by M2, wherein the k(cat) values for the derivatives MnM2-SgAP are different yet similar between MnCo- and CoCo-SgAP and pairs of other metal derivatives. On the other hand, BNPP hydrolysis is affected by metals in both sites. Thus, the two hydrolytic catalyses must follow different mechanisms. Based on crystal structures, docking, and the results presented herein, the M1 site is close to the hydrophobic specific site and the M2 site is next to Tyr246 that is H-bonded to a coordinated nucleophilic water molecule in peptide hydrolysis; whereas a coordinated water molecule on M1 becomes available as the nucleophile in phosphodiester hydrolysis.