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Catalytic Turnover (catalytic + turnover)
Selected AbstractsDifferential inhibition in vivo of ammonia monooxygenase, soluble methane monooxygenase and membrane-associated methane monooxygenase by phenylacetyleneENVIRONMENTAL MICROBIOLOGY, Issue 5 2000Sonny Lontoh Phenylacetylene was investigated as a differential inhibitor of ammonia monooxygenase (AMO), soluble methane monooxygenase (sMMO) and membrane-associated or particulate methane monooxygenase (pMMO) in vivo. At phenylacetylene concentrations >,1 µM, whole-cell AMO activity in Nitrosomonas europaea was completely inhibited. Phenylacetylene concentrations above 100 µM inhibited more than 90% of sMMO activity in Methylococcus capsulatus Bath and Methylosinus trichosporium OB3b. In contrast, activity of pMMO in M. trichosporium OB3b, M. capsulatus Bath, Methylomicrobium album BG8, Methylobacter marinus A45 and Methylomonas strain MN was still measurable at phenylacetylene concentrations up to 1000 µM. AMO of Nitrosococcus oceanus has more sequence similarity to pMMO than to AMO of N. europaea. Correspondingly, AMO in N. oceanus was also measurable in the presence of 1000 µM phenylacetylene. Measurement of oxygen uptake indicated that phenylacetylene acted as a specific and mechanistic-based inhibitor of whole-cell sMMO activity; inactivation of sMMO was irreversible, time dependent, first order and required catalytic turnover. Corresponding measurement of oxygen uptake in whole cells of methanotrophs expressing pMMO showed that pMMO activity was inhibited by phenylacetylene, but only if methane was already being oxidized, and then only at much higher concentrations of phenylacetylene and at lower rates compared with sMMO. As phenylacetylene has a high solubility and low volatility, it may prove to be useful for monitoring methanotrophic and nitrifying activity as well as identifying the form of MMO predominantly expressed in situ. [source] Substitution- and Elimination-Free Phosphorylation of Functionalized Alcohols Catalyzed by Oxidomolybdenum TetrachlorideADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 1 2010Cheng-Yuan Liu Abstract Among 14 oxidometallic species examined for catalytic phosphorylation of the tested alcohols, oxidomolybdenum tetrachloride (MoOCl4) was found to be the most efficient with a negligible background reaction mediated by triethylamine (Et3N). The new catalytic protocol can be applied to the chemoselective phosphorylations of primary, secondary and tertiary alcohols as well as the substitution-free phosphorylations of allylic, propargylic, and benzylic alcohols. Functionalized alcohols bearing acetonide, tetrahydropyranyl ether, tert -butyldimethylsilyl ether, or ester group are also amenable to the new catalytic protocol. The most difficult scenarios involve substitution-free phosphorylations of 1-phenylethanol and 1-(2-naphthyl)ethanol which can be effected in 95 and 90% yields, respectively. ESI-MS, IR, 1H, and 31P,NMR spectroscopic analyses of the reaction progress suggest the intermediacy of an alkoxyoxidomolybdenum trichloride-triethylamine adduct such as [(RO)Mo(O)Cl3 -Et3N] to be responsible for the catalytic turnover. [source] Osmium-Catalyzed Olefin Dihydroxylation and Aminohydroxylation in the Second Catalytic CycleADVANCED SYNTHESIS & CATALYSIS (PREVIOUSLY: JOURNAL FUER PRAKTISCHE CHEMIE), Issue 9 2006Peng Wu Abstract Two catalytic cycles operate in the osmium-catalyzed olefin dihydroxylation and aminohydroxylation. Slow hydrolysis of the Os(VI) monoglycolate (or monoazaglycolate in aminohydroxylation) intermediate often results in the addition of another molecule of olefin thereby shunting the catalysis into the second catalytic cycle. As a result, both enantio- and chemoselectivity are reduced. A series of new chelating ligands were devised, which force the catalysis into the second cycle while maintaining enantiocontrol in the olefin addition step. Excellent catalytic turnover and moderate to good enantioselectivity were achieved. [source] High tolerance to simultaneous active-site mutations in TEM-1 ,-lactamase: Distinct mutational paths provide more generalized ,-lactam recognitionPROTEIN SCIENCE, Issue 1 2009Pierre-Yves De Wals Abstract The diversity in substrate recognition spectra exhibited by various ,-lactamases can result from one or a few mutations in the active-site area. Using Escherichia coli TEM-1 ,-lactamase as a template that efficiently hydrolyses penicillins, we performed site-saturation mutagenesis simultaneously on two opposite faces of the active-site cavity. Residues 104 and 105 as well as 238, 240, and 244 were targeted to verify their combinatorial effects on substrate specificity and enzyme activity and to probe for cooperativity between these residues. Selection for hydrolysis of an extended-spectrum cephalosporin, cefotaxime (CTX), led to the identification of a variety of novel mutational combinations. In vivo survival assays and in vitro characterization demonstrated a general tendency toward increased CTX and decreased penicillin resistance. Although selection was undertaken with CTX, productive binding (KM) was improved for all substrates tested, including benzylpenicillin for which catalytic turnover (kcat) was reduced. This indicates broadened substrate specificity, resulting in more generalized (or less specialized) variants. In most variants, the G238S mutation largely accounted for the observed properties, with additional mutations acting in an additive fashion to enhance these properties. However, the most efficient variant did not harbor the mutation G238S but combined two neighboring mutations that acted synergistically, also providing a catalytic generalization. Our exploration of concurrent mutations illustrates the high tolerance of the TEM-1 active site to multiple simultaneous mutations and reveals two distinct mutational paths to substrate spectrum diversification. [source] Enantioselective Epoxidation of Terminal Alkenes to (R)- and (S)-Epoxides by Engineered Cytochromes P450 BM-3CHEMISTRY - A EUROPEAN JOURNAL, Issue 4 2006Takafumi Kubo Abstract Cytochrome P450 BM-3 from Bacillus megaterium was engineered for enantioselective epoxidation of simple terminal alkenes. Screening saturation mutagenesis libraries, in which mutations were introduced in the active site of an engineered P450, followed by recombination of beneficial mutations generated two P450 BM-3 variants that convert a range of terminal alkenes to either (R) - or (S) - epoxide (up to 83,% ee) with high catalytic turnovers (up to 1370) and high epoxidation selectivities (up to 95,%). A biocatalytic system using E. coli lysates containing P450 variants as the epoxidation catalysts and in vitro NADPH regeneration by the alcohol dehydrogenase from Thermoanaerobium brockii generates each of the epoxide enantiomers, without additional cofactor. [source] | ||||||||||