Accessing a synthetic FeIIIMnIV core to model biological heterobimetallic active sites.

Metalloproteins with dinuclear cores are known to bind and activate dioxygen, with a subclass of these proteins having active sites containing FeMn cofactors and activities ranging from long-range proton-coupled electron transfer (PCET) to post-translational peptide modification. While mechanistic studies propose that these metallocofactors access FeIIIMnIV intermediates, there is a dearth of related synthetic analogs. Herein, the first well-characterized synthetic FeIII-(µ-O)-MnIV complex is reported; this complex shows similar spectroscopic features as the catalytically competent FeIIIMnIV intermediate X found in Class Ic ribonucleotide reductase and demonstrates PCET function towards phenolic substrates. This complex is prepared from the oxidation of the isolable FeIII-(µ-O)-MnIII species, whose stepwise assembly is facilitated by a tripodal ligand containing phosphinic amido groups. Structural and spectroscopic studies found proton movement involving the FeIIIMnIII core, whereby the initial bridging hydroxido ligand is converted to an oxido ligand with concomitant protonation of one phosphinic amido group. This series of FeMn complexes allowed us to address factors that may dictate the preference of an active site for a heterobimetallic cofactor over one that is homobimetallic: comparisons of the redox properties of our FeMn complexes with those of the di-Fe analogs suggested that the relative thermodynamic ease of accessing an FeIIIMnIV core can play an important role in determining the metal ion composition when the key catalytic steps do not require an overly potent oxidant. Moreover, these complexes allowed us to demonstrate the effect of the hyperfine interaction from non-Fe nuclei on 57Fe Mössbauer spectra which is relevant to MnFe intermediates in proteins.

Tropidonotus nicobarensis Sclater, 1891 is a junior synonym of Thamnophis saurita (Linnaeus, 1766) (Squamata: Serpentes: Natricinae).

Tropidonotus nicobarensis Sclater, 1891 is an enigmatic snake currently referred to the natricine genus Hebius Thompson, 1913. No specimens have been recorded since the original discovery of the name-bearing type in the late 19th century, which was allegedly collected on Kamorta Island in India's Nicobar Archipelago. Recently, a second observation of this species was reported from Havelock Island (Swarajdweep) in the nearby Andaman Archipelago. However, the snake in question is clearly conspecific with another natricine, Thamnophis saurita (Linnaeus, 1766), native to eastern North America. This discovery prompted us to examine the type specimen of Tropidonotus nicobarensis, which revealed that it too is conspecific with Thamnophis saurita. The provenance of the Tropidonotus nicobarensis type specimen and the Havelock Island snake are discussed, with the latter likely representing an introduced animal.

Kukri snakes Oligodon Fitzinger, 1826 of the Western Palearctic with the resurrection of Contia transcaspica Nikolsky, 1902 (Reptilia, Squamata, Colubridae).

The kukri snakes of the genus Oligodon Fitzinger, 1826 reach the westernmost limits of their distribution in Middle and Southwest Asia (Afghanistan, Iran, and Turkmenistan), and the Palearctic portions of Pakistan. In this article, we review the systematics and distribution of the two species native to this region, Oligodon arnensis (Shaw, 1802) and Oligodon taeniolatus (Jerdon, 1853) based on an integrative approach combining morphological, molecular, and species distribution modeling (SDM) data. Phylogenetic analyses recover O. taeniolatus populations from Iran and Turkmenistan in a clade with the O. arnensis species complex, rendering the former species paraphyletic relative to O. taeniolatus sensu stricto on the Indian subcontinent. To correct this, we resurrect the name Contia transcaspica Nikolsky, 1902 from the synonymy of O. taeniolatus and assign it to populations in Middle-Southwest Asia. So far, Oligodon transcaspicus comb. et stat. nov. is known only from the Köpet-Dag Mountain Range of northeast Iran and southern Turkmenistan, but SDM mapping suggests it may have a wider range. Genetic samples of O. "arnensis" from northern Pakistan are nested in a clade sister to the recently described Oligodon churahensis Mirza, Bhardwaj & Patel, 2021, and are phylogenetically separate from O. arnensis sensu stricto in south India and Sri Lanka. Based on morphological similarity, the Afghanistan and Pakistan populations are assigned to Oligodon russelius (Daudin, 1803) and we synonymize O. churahensis with this species. Our investigation leads us to remove O. taeniolatus from the snake fauna of Afghanistan, Iran, and Turkmenistan, with the consequence that only Oligodon transcaspicus comb. et stat. nov. and O. russelius are present in these countries. Additional studies are needed to resolve the taxonomy of the O. taeniolatus and O. arnensis species complexes on the Indian subcontinent, and an updated key for both groups is provided.

Bioinspired Di-Fe Complexes: Correlating Structure and Proton Transfer over Four Oxidation States.

Metalloproteins with active sites containing di-Fe cores exhibit diverse chemical reactivity that is linked to the precise transfer of protons and electrons which directly involve the di-Fe units. The redox conversions are commonly corroborated by spectroscopic methods, but the associated structural changes are often difficult to assess, particularly those related to proton movements. This report describes the development of di-Fe complexes in which the movements of protons and electrons are pinpointed during the stepwise oxidation of a di-FeII species to one with an FeIIIFeIV core. Complex formation was promoted using the phosphinic amido tripodal ligand [poat]3- (N,N',N″-[nitrilotris(ethane-2,1-diyl)]tris(P,P-diphenylphosphinic amido)) that provided dynamic coordination spheres that assisted in regulating both electron and proton transfer processes. Oxidation of an [FeII-(µ-OH)-FeIII] complex led to the corresponding di-FeIII species containing a hydroxido bridge that was not stable at room temperature and converted to a species containing an oxido bridging ligand and protonation of one phosphinic amido group to form [Hpoat]2-. Deprotonation led to a new species with an [FeIII-(µ-O)-FeIII] core that could be further oxidized to its FeIIIFeIV analogue. Reactions with phenols suggest homolytic cleavage of the O-H bond to give products that are consistent with the initial formation of a phenoxyl radical─spectroscopic studies indicated that the electron is transferred to the FeIV center, and the proton is initially transferred to the more sterically hindered oxido ligand but then relocates to [poat]3-. These findings offer new mechanistic insights related to the stability of and the reactions performed by di-Fe enzymes.

C-H Bond Cleavage by Bioinspired Nonheme Metal Complexes.

The functionalization of C-H bonds is one of the most challenging transformations in synthetic chemistry. In biology, these processes are well-known and are achieved with a variety of metalloenzymes, many of which contain a single metal center within their active sites. The most well studied are those with Fe centers, and the emerging experimental data show that high-valent iron oxido species are the intermediates responsible for cleaving the C-H bond. This Forum Article describes the state of this field with an emphasis on nonheme Fe enzymes and current experimental results that provide insights into the properties that make these species capable of C-H bond cleavage. These parameters are also briefly considered in regard to manganese oxido complexes and Cu-containing metalloenzymes. Synthetic iron oxido complexes are discussed to highlight their utility as spectroscopic and mechanistic probes and reagents for C-H bond functionalization. Avenues for future research are also examined.

Stepwise assembly of heterobimetallic complexes: synthesis, structure, and physical properties.

Bimetallic active sites are ubiquitous in metalloenzymes and have sparked investigations of synthetic models to aid in the establishment of structure-function relationship. We previously reported a series of discrete bimetallic complexes with [FeIII-(µ-OH)-MII] cores in which the ligand framework provides distinct binding sites for two metal centers. The formation of these complexes relied on a stepwise synthetic approach in which an FeIII-OH complex containing a sulfonamido tripodal ligand served as a synthon that promoted assembly. We have utilized this approach in the present study to produce a new series of bimetallic complexes with [FeIII-(µ-OH)-MII] cores (M = Ni, Cu, Zn) by using an ancillary ligand to the FeIII center that contains phosphinic amido groups. Assembly began with formation of an FeIII-OH that was subsequently used to bind the MII fragment that contained a triazacyclononane ligand. The series of bimetallic complexes were charactered structurally by X-ray diffraction methods, spectroscopically by absorption, vibrational, electron paramagnetic resonance spectroscopies, and electrochemically by cyclic voltammetry. A notable finding is that these new [FeIII-(µ-OH)-MII] complexes displayed significantly lower reduction potentials than their sulfonamido counterparts, which paves way for future studies on high valent bimetallic complexes in this scaffold.

CowN sustains nitrogenase turnover in the presence of the inhibitor carbon monoxide.

Nitrogenase is the only enzyme capable of catalyzing nitrogen fixation, the reduction of dinitrogen gas (N2) to ammonia (NH3). Nitrogenase is tightly inhibited by the environmental gas carbon monoxide (CO). Nitrogen-fixing bacteria rely on the protein CowN to grow in the presence of CO. However, the mechanism by which CowN operates is unknown. Here, we present the biochemical characterization of CowN and examine how CowN protects nitrogenase from CO. We determine that CowN interacts directly with nitrogenase and that CowN protection observes hyperbolic kinetics with respect to CowN concentration. At a CO concentration of 0.001 atm, CowN restores nearly full nitrogenase activity. Our results further indicate that CowN's protection mechanism involves decreasing the binding affinity of CO to nitrogenase's active site approximately tenfold without interrupting substrate turnover. Taken together, our work suggests CowN is an important auxiliary protein in nitrogen fixation that engenders CO tolerance to nitrogenase.

Lignin-Derived Non-Heme Iron and Manganese Complexes: Catalysts for the On-Demand Production of Chlorine Dioxide in Water under Mild Conditions.

A lignin-derived ligand, bis(2-hydroxy-3-methoxy-5-propylbenzyl)glycine (DHEG), was synthesized from 2-methoxy-4-propylphenol (dihydroeugenol (DHE)) and the amino acid glycine. Two mononuclear iron and manganese complexes of DHEG were prepared, characterized, and employed for the oxidation of chlorite to chlorine dioxide in aqueous solution. Peroxyacetic acid (PAA) was used as a "green" oxidant in the redox reactions to initiate the formation of high-valent Fe and Mn (IV)-OH intermediates. EPR studies verified the formation of a high-valent MnIV species. Both Fe and Mn complexes catalyzed chlorite oxidation with bimolecular rate constants of 32 and 144 M-1 s-1, respectively, at pH 4.0 and 25 °C. The Mn complex was found to be more efficient for chlorite oxidation with a turnover frequency of 17 h-1 and remained active during subsequent additions of PAA. The rate of ClO2 decomposition with PAA/Mn-DHEG was first order in PAA and increased significantly as pH increased. A mechanism that accounts for all observations is presented.

Effects of Noncovalent Interactions on High-Spin Fe(IV)-Oxido Complexes.

High-valent nonheme FeIV-oxido species are key intermediates in biological oxidation, and their properties are proposed to be influenced by the unique microenvironments present in protein active sites. Microenvironments are regulated by noncovalent interactions, such as hydrogen bonds (H-bonds) and electrostatic interactions; however, there is little quantitative information about how these interactions affect crucial properties of high valent metal-oxido complexes. To address this knowledge gap, we introduced a series of FeIV-oxido complexes that have the same S = 2 spin ground state as those found in nature and then systematically probed the effects of noncovalent interactions on their electronic, structural, and vibrational properties. The key design feature that provides access to these complexes is the new tripodal ligand [poat]3-, which contains phosphinic amido groups. An important structural aspect of [FeIVpoat(O)]- is the inclusion of an auxiliary site capable of binding a Lewis acid (LAII); we used this unique feature to further modulate the electrostatic environment around the Fe-oxido unit. Experimentally, studies confirmed that H-bonds and LAII s can interact directly with the oxido ligand in FeIV-oxido complexes, which weakens the Fe═O bond and has an impact on the electronic structure. We found that relatively large vibrational changes in the Fe-oxido unit correlate with small structural changes that could be difficult to measure, especially within a protein active site. Our work demonstrates the important role of noncovalent interactions on the properties of metal complexes, and that these interactions need to be considered when developing effective oxidants.

Epoxidation Catalyzed by the Nonheme Iron(II)- and 2-Oxoglutarate-Dependent Oxygenase, AsqJ: Mechanistic Elucidation of Oxygen Atom Transfer by a Ferryl Intermediate.

Mechanisms of enzymatic epoxidation via oxygen atom transfer (OAT) to an olefin moiety is mainly derived from the studies on thiolate-heme containing epoxidases, such as cytochrome P450 epoxidases. The molecular basis of epoxidation catalyzed by nonheme-iron enzymes is much less explored. Herein, we present a detailed study on epoxidation catalyzed by the nonheme iron(II)- and 2-oxoglutarate-dependent (Fe/2OG) oxygenase, AsqJ. The native substrate and analogues with different para substituents ranging from electron-donating groups (e.g., methoxy) to electron-withdrawing groups (e.g., trifluoromethyl) were used to probe the mechanism. The results derived from transient-state enzyme kinetics, Mössbauer spectroscopy, reaction product analysis, X-ray crystallography, density functional theory calculations, and molecular dynamic simulations collectively revealed the following mechanistic insights: (1) The rapid O2 addition to the AsqJ Fe(II) center occurs with the iron-bound 2OG adopting an online-binding mode in which the C1 carboxylate group of 2OG is trans to the proximal histidine (His134) of the 2-His-1-carboxylate facial triad, instead of assuming the offline-binding mode with the C1 carboxylate group trans to the distal histidine (His211); (2) The decay rate constant of the ferryl intermediate is not strongly affected by the nature of the para substituents of the substrate during the OAT step, a reactivity behavior that is drastically different from nonheme Fe(IV)-oxo synthetic model complexes; (3) The OAT step most likely proceeds through a stepwise process with the initial formation of a C(benzylic)-O bond to generate an Fe-alkoxide species, which is observed in the AsqJ crystal structure. The subsequent C3-O bond formation completes the epoxide installation.

The discovery of Rock Geckos Cnemaspis Strauch, 1887 (Squamata: Gekkonidae) in the Tanintharyi Region, Myanmar with the description of two new species.

We report the first occurrence of the Asian Rock Gecko genus Cnemaspis Strauch, 1887 from mainland Myanmar based on a series of specimens recently collected from the Tanintharyi Region. These records come from several localities in the Tanintharyi Region and fill a significant sampling gap for the genus. Molecular phylogenetic analyses using the mitochondrial gene ND2 identify two distinct clades, the first includes Cnemaspis siamensis of the Southeast Asian Cnemaspis group and the second includes two new species belonging to the South Asian Cnemaspis kandiana group. These two species are morphologically distinct and are distinguished from all other members in the C. kandiana group by a combination of character states. The first species, Cnemaspis tanintharyi sp. nov. occurs on the mainland in southern Tanintharyi and is distinguished from all congeners by the possession of keeled pectoral scales; smooth ventral scales and abdominal scales; 2-4 precloacal pores; 4-5 femoral pores on each leg; smooth subcaudal scales with the median row enlarged; coloration of the gular region beige, dark gray-brown with dark blue hueing towards throat; 15-18 subdigital lamellae on the 4th toe; and 21-23 ventral scales at midbody. The other, Cnemaspis thayawthadangyi sp. nov. is known only from the island group of Thayawthadangyi, in the Myeik Archipelago and is distinguished from all congeners by the possession of keeled pectoral scales; keeled ventral scales and abdominal scales; three precloacal pores; four femoral pores on each leg; smooth subcaudal scales, scales on median row enlarged; coloration of the gular region silver with dark-gray irregularly shaped streaks; 16-18 subdigital lamellae on the 4th toe; and 18-20 ventral scales at midbody. In addition, we address the taxonomic status of populations referred to as C. kandiana (a species now restricted to Sri Lanka) found in peninsular Thailand and provisionally assign them as C. cf. tanintharyi sp. nov. until further analysis can be conducted. Finally, we briefly discuss the biogeography of the South Asian clade of Cnemaspis.

A global catalog of primary reptile type specimens.

We present information on primary type specimens for 13,282 species and subspecies of reptiles compiled in the Reptile Database, that is, holotypes, neotypes, lectotypes, and syntypes. These represent 99.4% of all 13,361 currently recognized taxa (11,050 species and 2311 subspecies). Type specimens of 653 taxa (4.9%) are either lost or not located, were never designated, or we did not find any information about them. 51 species are based on iconotypes. To map all types to physical collections we have consolidated all synonymous and ambiguous collection acronyms into an unambiguous list of 364 collections holding these primary types. The 10 largest collections possess more than 50% of all (primary) reptile types, the 36 largest collections possess more than 10,000 types and the largest 73 collections possess over 90% of all types. Of the 364 collections, 107 hold type specimens of only 1 species or subspecies. Dozens of types are still in private collections. In order to increase their utility, we recommend that the description of type specimens be supplemented with data from high-resolution images and CT-scans, and clear links to tissue samples and DNA sequence data (when available). We request members of the herpetological community provide us with any missing type information to complete the list.

Filling the BINs of life: Report of an amphibian and reptile survey of the Tanintharyi (Tenasserim) Region of Myanmar, with DNA barcode data.

Despite threats of species extinctions, taxonomic crises, and technological advances in genomics and natural history database informatics, we are still distant from cataloguing all of the species of life on earth. Amphibians and reptiles are no exceptions; in fact new species are described nearly every day and many species face possible extinction. The number of described species continues to climb as new areas of the world are explored and as species complexes are examined more thoroughly. The use of DNA barcoding provides a mechanism for rapidly estimating the number of species at a given site and has the potential to record all of the species of life on Earth. Though DNA barcoding has its caveats, it can be useful to estimate the number of species in a more systematic and efficient manner, to be followed in combination with more traditional, morphology-based identifications and species descriptions. Herein, we report the results of a voucher-based herpetological expedition to the Tanintharyi (Tenasserim) Region of Myanmar, enhanced with DNA barcode data. Our main surveys took place in the currently proposed Tanintharyi National Park. We combine our results with photographs and observational data from the Chaung-nauk-pyan forest reserve. Additionally, we provide the first checklist of amphibians and reptiles of the region, with species based on the literature and museum. Amphibians, anurans in particular, are one of the most poorly known groups of vertebrates in terms of taxonomy and the number of known species, particularly in Southeast Asia. Our rapid-assessment program combined with DNA barcoding and use of Barcode Index Numbers (BINs) of voucher specimens reveals the depth of taxonomic diversity in the southern Tanintharyi herpetofauna even though only a third of the potential amphibians and reptiles were seen. A total of 51 putative species (one caecilian, 25 frogs, 13 lizards, 10 snakes, and two turtles) were detected, several of which represent potentially undescribed species. Several of these species were detected by DNA barcode data alone. Furthermore, five species were recorded for the first time in Myanmar, two amphibians (Ichthyophis cf. kohtaoensis and Chalcorana eschatia) and three snakes (Ahaetulla mycterizans, Boiga dendrophila, and Boiga drapiezii).

Insights into the Desaturation of Cyclopeptin and its C3 Epimer Catalyzed by a non-Heme Iron Enzyme: Structural Characterization and Mechanism Elucidation.

AsqJ, an iron(II)- and 2-oxoglutarate-dependent enzyme found in viridicatin-type alkaloid biosynthetic pathways, catalyzes sequential desaturation and epoxidation to produce cyclopenins. Crystal structures of AsqJ bound to cyclopeptin and its C3 epimer are reported. Meanwhile, a detailed mechanistic study was carried out to decipher the desaturation mechanism. These findings suggest that a pathway involving hydrogen atom abstraction at the C10 position of the substrate by a short-lived FeIV -oxo species and the subsequent formation of a carbocation or a hydroxylated intermediate is preferred during AsqJ-catalyzed desaturation.

Troublesome Trimes: Potential cryptic speciation of the Trimeresurus (Popeia) popeiorum complex (Serpentes: Crotalidae) around the Isthmus of Kra (Myanmar and Thailand).

The taxonomic identity of the Trimeresurus (Popeia) popeiorum complex from the Isthmus of Kra and to the north was investigated. Several studies over the last decade have produced several specimens and associated mtDNA sequence data for a variety of individuals of the T. popeiorum and "T. sabahi" complexes. Here, we combine four mitochondrial genes (12S, 16S, ND4, and CytB) from all available specimens in GenBank with the addition of five new specimens collected from the mainland, Tanintharyi Region of Myanmar. Maximum Likelihood and Bayesian analyses identified that T. popeiorum sensu lato is paraphyletic with two geographically distinct clades: a northern clade representing populations from northern Myanmar, Laos and northern Thailand and a southern clade representing samples from the Tanintharyi Region and adjacent west Thailand. While the two clades have considerable genetic distance, they appear to be morphologically identical, leading to the hypothesis that the southern clade represents a cryptic, undescribed species. Because they appear to be cryptic species and the limitation of only five specimens from the southern lineage, this does not permit us to formally describe the new species. In accordance to past molecular studies, we uncovered paraphyly and lack of genetic support for the validity of taxa within the T. sabahi complex. However, we suggest recognizing these populations as subspecies within T. sabahi.

Mechanistic Investigation of a Non-Heme Iron Enzyme Catalyzed Epoxidation in (-)-4'-Methoxycyclopenin Biosynthesis.

Mechanisms have been proposed for α-KG-dependent non-heme iron enzyme catalyzed oxygen atom insertion into an olefinic moiety in various natural products, but they have not been examined in detail. Using a combination of methods including transient kinetics, Mössbauer spectroscopy, and mass spectrometry, we demonstrate that AsqJ-catalyzed (-)-4'-methoxycyclopenin formation uses a high-spin Fe(IV)-oxo intermediate to carry out epoxidation. Furthermore, product analysis on (16)O/(18)O isotope incorporation from the reactions using the native substrate, 4'-methoxydehydrocyclopeptin, and a mechanistic probe, dehydrocyclopeptin, reveals evidence supporting oxo↔hydroxo tautomerism of the Fe(IV)-oxo species in the non-heme iron enzyme catalysis.