The particulate methane monooxygenase (pMMO) is known to be very difficult to study mainly due to its unusual activity instability in vitro. By cultivating Methylococcus capsulatus (Bath) under methane stress conditions and high copper levels in the growth medium, membranes highly enriched in the pMMO with exceptionally stable activity can be isolated from these cells. Purified and active pMMO can be subsequently obtained from these membrane preparations using protocols in which an excess of reductants and anaerobic conditions were maintained during membrane solubilization by dodecyl ␤-D-maltoside and purification by chromatography. The pMMO was found to be the major constituent in these membranes, constituting 60 -80% of total membrane proteins. The dominant species of the pMMO was found to consist of three subunits, ␣, ␤, and ␥, with an apparent molecular mass of 45, 26, and 23 kDa, respectively. A second species of the pMMO, a proteolytically processed version of the enzyme, was found to be composed of three subunits, ␣, ␤, and ␥, with an apparent molecular mass of 35, 26, and 23 kDa, respectively. The ␣ and ␣ subunits from these two forms of the pMMO contain identical N-terminal sequences. The ␥ subunit, however, exhibits variation in its N-terminal sequence. The pMMO is a copper-containing protein only and shows a requirement for Cu(I) ions. Approximately 12-15 Cu ions per 94-kDa monomeric unit were observed. The pMMO is sensitive to dioxygen tension. On the basis of dioxygen sensitivity, three kinetically distinct forms of the enzyme can be distinguished. A slow but air-stable form, which is converted into a "pulsed" state upon direct exposure to atmospheric oxygen pressure, is considered as type I pMMO. This form was the subject of our pMMO isolation effort. Other forms (types II and III) are deactivated to various extents upon exposure to atmospheric dioxygen pressure. Under inactivating conditions, these unstable forms release protons to the buffer (ϳ10 H ؉ /94-kDa monomeric unit) and eventually become completely inactive.The enzyme methane monooxygenase, found in methanotrophic bacteria, catalyzes the conversion of methane to methanol using dioxygen as a co-substrate at ambient temperatures and pressures (1, 2). This system has attracted considerable attention, since it provides an ideal natural model to study methane activation and functionalization, a subject of significant current interest (3). Two distinct species of methane monooxygenase (MMO) 1 are known to exist at different cellular locations, a cytoplasmic (soluble) MMO and a membrane-bound (particulate) MMO (4). The soluble MMO (sMMO) is a complex threecomponent system consisting of a hydroxylase, a reductase, and a small regulatory protein (4). The sMMO has been investigated extensively by several research groups (5-21). The xray crystal structure of the sMMO hydroxylase isolated from Methylococcus capsulatus (Bath) has been solved (22, 23). The hydroxylase active site contains a non-heme binuclear iron cluster. In contrast, the particulat...

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The regiospecificity and stereoselectivity of alkane hydroxylation and alkene epoxidation by the particulate methane monooxygenase from Methylococcus capsulatus (Bath) was evaluated over a range of substrates. Oxidation products were identified by conventional GC analysis, and the stereoselectivity of oxidation was determined by a combination of chiral GC and HPLC methods, as well as 1H NMR analysis of the corresponding (R)-2-acetoxy-2-phenylethanoate ester derivatives in the case of alkanol products. Alkane hydroxylation was found to proceed favoring attack at the C-2 position in all cases, and the stereoselectivity for n-butane and n-pentane was characterized by an enantiomeric excess of 46% and 80%, respectively, with preference for the (R)-alcohol noted for both substrates. Epoxides were formed with smaller stereoselectivities. Together, the regio- and stereoselectivity results suggest that an equilibrium of competing substrate binding modes exists. A simple substrate-binding model that incorporates preferential C-2 oxidation with the observed stereoselectivity of alkane hydroxylation is proposed, and hypotheses for the general mechanism are suggested and discussed.

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A series of pyrenyl complexes containing one or two AuPPh3 or Pt(PEt3)2Br groups have been synthesized to probe the effects of metal on the absorption and emission properties of the organic molecule. The absorption spectra of the complexes showed perturbation of the metal centers on the pyrenyl ring, as manifested by a red shift of the π → π* transitions of pyrene and intensification of the forbidden 1Lb band. The perturbation increased with the number of metal ions attached to the pyrenyl ring. The Pt ion was stronger than the Au ion in perturbing the electronic structure of the pyrenyl ring. All pyrenyl complexes displayed fluorescence and phosphorescence in degassed solutions. The heavy-atom effect of the metal ions enhanced the phosphorescence. The quantum yield of phosphorescence was sensitive to the positions of metalation.

A luminescent molecular rectangle [Au(4)(micro-PAnP)(2)(micro-bipy)(2)](OTf)(4) (1.(OTf)(4)) (PAnP = 9,10-bis(diphenylphosphino)anthracene, bipy = 4,4'-bipyridine, X = NO(3)(-) or OTf(-)), synthesized from the self-assembly of the molecular "clip" Au(2)(micro-PAnP)(OTf)(2) and bipy, shows a large rectangular cavity of 7.921(3) x 16.76(3) A. The electronic absorption and emission spectroscopy, and electrochemistry of the metallacyclophane, have been studied. The 1(4+) ions are self-assembled into 2D mosaic in the solid state via complementary edge-to-face interactions between the Ph groups. (1)H NMR titrations ratify the 1:1 complexation between 1(4+) and various aromatic molecules. Comparing the structures of the inclusion complexes indicates an induced-fit mechanism operating in the binding. The emission of 1(4+) is quenched upon the guest binding. The binding constants are determined by both (1)H NMR and fluorescence titrations. Solvophobic and ion-dipole effects are shown to be important in stabilizing the inclusion complexes.

A new anionic heteroleptic Ir(III)-dithiolate complex Ir(ppy)2(benzene-1,2-dithiolate) (ppy = 2-phenylpyridine, [IrSS](-)) undergoes very fast air oxidation to form a monosulfinate complex [IrSSO2](-), which can be further dioxygenated by O2 or H2O2 to give a disulfinate complex [IrSO2SO2](-), which has been characterized by X-ray crystallography. The dioxygenation is accompanied by changes in the electronic structures of the complexes, leading to blue shift of emission from [IrSS](-) (λ(max) = 665 nm) to [IrSSO2](-) (λ(max) = 556 nm) and to [IrSO2SO2](-) (λ(max) = 460 nm). The molecular and electronic structures of the complexes are probed by DFT calculations. Time-dependent DFT (TD-DFT) calculations show the lowest energy spin-allowed electronic transitions for [IrSS](-) and [IrSSO2](-) are mainly ligand (3p orbital of S)-to-ligand (π* orbitals of ppy)-charge-transfer transition, whereas the lowest energy electronic transition in [IrSO2SO2](-) is predominantly metal-to-ligand-charge transfer in nature.

A n e w luminescent d8-d8 dinuclear Ptll polypyridyl complex [Pt2(2,2' : 6',2''-terp~ridine)~(Gua)]3+ is synthesised and characterized by X-ray crystallography; its spectroscopic and photoredox properties are investigated.