Enhancing predictive power by unamalgamating multi-item scales.

The generally small but touted as "statistically significant" correlation coefficients in the social sciences jeopardize theory testing and prediction. To investigate these small coefficients' underlying causes, traditional equations such as Spearman's (1904) classic attenuation formula, Cronbach's (1951) alpha, and Guilford and Fruchter's (1973) equation for the effect of additional items on a scale's predictive power are considered. These equations' implications differ regarding large interitem correlations enhancing or diminishing predictive power. Contrary to conventional practice, such correlations decrease predictive power when treating items as multi-item scale components but can increase predictive power when treating items separately. The implications are wide-ranging. (PsycInfo Database Record (c) 2023 APA, all rights reserved).

Diyne inactivators and activity-based fluorescent labeling of phenol hydroxylase in Pseudomonas sp. CF600.

An activity-based labeling (ABL) approach was investigated for the phenol-oxidizing bacterium, Pseudomonas sp. CF600. Phenol-grown cells were exposed to several different terminal diynes, and following cell breakage, extracts of these cells were added to copper-catalyzed alkyne/azide cycloaddition reactions containing Alexa Fluor 647 azide. Analysis of total cell proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and near-infrared scanning demonstrated covalent fluorescent labeling of a 58- and a 34-kDa polypeptide in all diyne-treated cell types. Further studies using 1,4-diethynylbenzene (DEB) demonstrated that these labeled polypeptides were consistently detected in cells grown on substrates that exhibited phenol-dependent O2 uptake activity but not observed when cells were grown on substrates such as dextrose or catechol that did not support this activity. Fluorescent labeling of the two polypeptides in DEB-treated, phenol-grown cells was time dependent and was inhibited by several known substrates for phenol hydroxylase. These results suggest that diverse diynes act as mechanism-based inactivators of phenol hydroxylase in Pseudomonas sp. CF600 and that this effect can be exploited by ABL approaches to selectively label the major 58- and 34-kDa subunits of the hydroxylase component of this complex enzyme.

Pilot-scale biofiltration of 1,4-dioxane at drinking water-relevant concentrations.

1,4-Dioxane is a drinking water contaminant of emerging concern. Because conventional and many advanced drinking water treatment technologies are ineffective for 1,4-dioxane removal, cost-effective technologies for the removal of 1,4-dioxane at drinking water-relevant concentrations are needed. In this research, a gravity-fed, cometabolic biofiltration system was developed to degrade 1,4-dioxane that was spiked into coagulated, settled surface water at a concentration of ∼10 µg/L. Objectives were to determine whether cometabolic degradation of trace levels of 1,4-dioxane can be sustained using n-butane as primary substrate and whether filter media properties and empty bed contact time (EBCT) affect biofiltration efficiency. A mixed culture of bacteria derived from the Cape Fear River basin and previously enriched using isobutane served as inoculum for biologically active filters. Two granular activated carbons (GACs) with different grain sizes and one carbonaceous resin were used as attachment media, and n-butane served as the primary substrate for biologically active filters. Non-inoculated controls with the same media were evaluated in parallel to distinguish between biological and adsorptive removals of 1,4-dioxane. For the duration of the pilot study (>3 months), 1,4-dioxane was degraded in inoculated biofilters receiving n-butane. In control filters containing larger and smaller grain GAC, 1,4-dioxane broke through completely within 750 and 1250 bed volumes, respectively, corresponding to 15 to 30 days of operation at an EBCT of 30 min. 1,4-Dioxane removal increased with increasing EBCT in all biologically active filters. At an EBCT of 30 min, the biologically active GAC filter containing the larger-grain GAC removed on average 87% of 1,4-dioxane at pseudo steady-state. When the hydraulic loading rate was decreased to achieve an overall EBCT of 60 min, 1,4-dioxane was removed to <1 µg/L in the biologically active GAC filter containing the larger-grain GAC. Activity-based labeling showed the presence of catalytically active monooxygenases in backwash water from biologically active filters that degraded 1,4-dioxane. Amplicon sequencing results showed that while taxa shifted after the initial inoculation of biologically active filters, taxa in biologically active filters remained more similar to the inoculum than those in the non-inoculated control filters. Overall, results of this research demonstrate that cometabolic degradation of 1,4-dioxane at trace levels is possible for extended periods of time in inoculated biofilters that receive n-butane as primary substrate.

The Ethics and Politics of Academic Knowledge Production: Thoughts on the Future of Business Ethics.

To commemorate 40 years since the founding of the Journal of Business Ethics, the editors in chief of the journal have invited the editors to provide commentaries on the future of business ethics. This essay comprises a selection of commentaries aimed at creating dialogue around the theme The Ethics and Politics of Academic Knowledge Production. Questions of who produces knowledge about what, and how that knowledge is produced, are inherent to editing and publishing academic journals. At the Journal of Business Ethics, we understand the ethical responsibility of academic knowledge production as going far beyond conventions around the integrity of the research content and research processes. We are deeply aware that access to resources, knowledge of the rules of the game, and being able to set those rules, are systematically and unequally distributed. One could ask the question "for whom is knowledge now ethical'"? (See the Burrell commentary.) We have a responsibility to address these inequalities and open up our journal to lesser heard voices, ideas, and ways of being. Our six commentators pursue this through various aspects of the ethics and politics of academic knowledge production. Working with MacIntyre's scheme of practices and institutions, Andrew West provides commentary on the internal good of business ethics learning and education. Inviting us to step out of the cave, Christopher Michaelson urges a clear-eyed, unblinking focus on the purposes and audiences of business ethics scholarship. As developmental editor, Scott Taylor uncovers some of the politics of peer review with the aim of nurturing of unconventional research. Mike Hyman presents his idiosyncratic view of marketing ethics. In the penultimate commentary, Julie Nelson attributes difficulties in the academic positioning of the Business Ethics field to the hegemony of a masculine-centric model of the firm. And finally, Gibson Burrell provides a powerful provocation to go undercover as researcher-investigators in a parallel ethics of the research process.

Co-encapsulation of slow release compounds and Rhodococcus rhodochrous ATCC 21198 in gellan gum beads to promote the long-term aerobic cometabolic transformation of 1,1,1-trichloroethane, cis-1,2-dichloroethene and 1,4-dioxane.

Rhodococcus rhodochrous ATCC 21198 (strain ATCC 21198) was successfully co-encapsulated in gellan gum beads with orthosilicates as slow release compounds (SRCs) to support aerobic cometabolism of a mixture of 1,1,1-trichloroethane (1,1,1-TCA), cis-1,2-dichloroethene (cis-DCE), and 1,4-dioxane (1,4-D) at aqueous concentrations ranging from 250 to 1000 µg L-1. Oxygen (O2) consumption and carbon dioxide (CO2) production showed the co-encapsulated cells utilized the alcohols that were released from the co-encapsulated SRCs. Two model SRCs, tetrabutylorthosilicate (TBOS) and tetra-s-butylorthosilicate (T2BOS), which hydrolyze to produce 1- and 2-butanol, respectively, were encapsulated in gellan gum (GG) at mass loadings as high as 10% (w/w), along with strain ATCC 21198. In the GG encapsulated beads, TBOS hydrolyzed 26 times faster than T2BOS and rates were ∼4 times higher in suspension than when encapsulated. In biologically active reactors, the co-encapsulated strain ATCC 21198 effectively utilized the SRC hydrolysis products (1- and 2-butanol) and cometabolized repeated additions of a mixture of 1,1,1-TCA, cis-DCE, and 1,4-D for over 300 days. The transformation followed pseudo-first-order kinetics. Vinyl chloride (VC) and 1,1-dichloroethene (1,1-DCE) were also transformed in the reactors after 250 days. In the long-term treatment, the batch reactors with co-encapsulated T2BOS GG beads achieved similar transformation rates, but at much lower O2 consumption rates than those with TBOS. The results demonstrate that the co-encapsulation technology can be a passive method for the cometabolic treatment of dilute groundwater plumes.

Aerobic cometabolism of 1,4-dioxane by isobutane-utilizing microorganisms including Rhodococcus rhodochrous strain 21198 in aquifer microcosms: Experimental and modeling study.

Aerobic cometabolism of the emerging contaminant 1,4-dioxane (1,4-D) by isobutane-utilizing microorganisms was assessed in pure culture and aquifer microcosm studies. The bacterium Rhodococcus rhodochrous strain ATCC 21198 transformed low, environmentally-relevant concentrations of 1,4-D when grown on isobutane. Microcosms were constructed with aquifer solids from Fort Carson, Colorado, a site contaminated with 1,4-D and trichloroethene (TCE). Multiple additions of isobutane and 1,4-D over 300 days were transformed in microcosms biostimulated with isobutane and microcosms bioaugmented with strain 21198. Results showed that, over time and with sufficient inorganic nutrients, biostimulation of native microorganisms with isobutane was just as effective as bioaugmentation with strain 21198 to achieve 1,4-D transformation in the microcosms. The presence of TCE at 0.2 mg/L did not inhibit 1,4-D transformation, though TCE itself was not readily transformed. An iterative process was used to determine kinetic parameter values to fit Michaelis-Menten/Monod models to experimental data for simultaneous isobutane utilization, biomass growth, and cometabolic transformation of 1,4-D. Parameter optimization resulted in good model fit to the data over multiple transformations of isobutane and 1,4-D in both short- and long-term experiments. Results suggest low concentrations of 1,4-D studied in the microcosms were cometabolically transformed according to a pseudo first-order rate of 0.37 L/mg TSS/day of 21198. Isobutane consumption was modeled with a maximum rate of 2.58 mg/mg TSS/day and a half saturation constant of 0.09 mg/L. 1,4-D transformation was competitively inhibited by the presence of isobutane and transformation rates were significantly reduced when inorganic nutrients were limiting. Simulations of the repeated additions found a first-order microbial endogenous decay coefficient of 0.03 day-1 fit the alternating periods of active transformation and stagnation between isobutane and 1,4-D additions over approximately one year. The model fitting process highlighted the importance of determining kinetic parameters from data representing low concentrations typically found in the environment.

Activity-Based Protein Profiling of Ammonia Monooxygenase in Nitrosomonas europaea.

Nitrosomonas europaea is an aerobic nitrifying bacterium that oxidizes ammonia (NH3) to nitrite (NO2 (-)) through the sequential activities of ammonia monooxygenase (AMO) and hydroxylamine dehydrogenase (HAO). Many alkynes are mechanism-based inactivators of AMO, and here we describe an activity-based protein profiling method for this enzyme using 1,7-octadiyne (17OD) as a probe. Inactivation of NH4 (+)-dependent O2 uptake by N. europaea by 17OD was time- and concentration-dependent. The effects of 17OD were specific for ammonia-oxidizing activity, andde novoprotein synthesis was required to reestablish this activity after cells were exposed to 17OD. Cells were reacted with Alexa Fluor 647 azide using a copper-catalyzed azide-alkyne cycloaddition (CuAAC) (click) reaction, solubilized, and analyzed by SDS-PAGE and infrared (IR) scanning. A fluorescent 28-kDa polypeptide was observed for cells previously exposed to 17OD but not for cells treated with either allylthiourea or acetylene prior to exposure to 17OD or for cells not previously exposed to 17OD. The fluorescent polypeptide was membrane associated and aggregated when heated with ß-mercaptoethanol and SDS. The fluorescent polypeptide was also detected in cells pretreated with other diynes, but not in cells pretreated with structural homologs containing a single ethynyl functional group. The membrane fraction from 17OD-treated cells was conjugated with biotin-azide and solubilized in SDS. Streptavidin affinity-purified polypeptides were on-bead trypsin-digested, and amino acid sequences of the peptide fragments were determined by liquid chromatography-mass spectrometry (LC-MS) analysis. Peptide fragments from AmoA were the predominant peptides detected in 17OD-treated samples. In-gel digestion and matrix-assisted laser desorption ionization-tandem time of flight (MALDI-TOF/TOF) analyses also confirmed that the fluorescent 28-kDa polypeptide was AmoA.

RNAi silencing of a cytochrome P450 monoxygenase disrupts the ability of a filamentous fungus, Graphium sp., to grow on short-chain gaseous alkanes and ethers.

Graphium sp. (ATCC 58400), a filamentous fungus, is one of the few eukaryotes that grows on short-chain alkanes and ethers. In this study, we investigated the genetic underpinnings that enable this fungus to catalyze the first step in the alkane and ether oxidation pathway. A gene, CYP52L1, was identified, cloned and functionally characterized as an alkane-oxidizing cytochrome P450 (GSPALK1). Analysis of CYP52L1 suggests that it is a member of the CYP52 cytochrome P450 family, which is comprised of medium- and long-chain alkane-oxidizing enzymes found in yeasts. However, phylogenetic analysis of GSPALK1 with other CYP52 members suggests they are not closely related. Post-transcriptional ds-RNA-mediated gene silencing of CYP52L1 severely reduced the ability of this fungus to oxidize alkanes and ethers, however, downstream metabolic steps in these pathways were unaffected. Collectively, the results of this study suggest that GSPALK1 is the enzyme that catalyzes the initial oxidation of alkanes and ethers but is not involved in the later steps of alkane or ether metabolism.

Oxidation of gasoline oxygenates by closely related non-haem-iron alkane hydroxylases in Pseudomonas mendocina KR1 and other n-octane-utilizing Pseudomonas strains.

Pseudomonas mendocina KR1 oxidizes the gasoline oxygenate methyl tertiary butyl ether (MTBE) to tertiary butyl alcohol (TBA) during growth on C5 -C8 n-alkanes. We have further explored oxidation of ether oxygenates by this strain to help identify the enzyme that catalyses these reactions. High levels of MTBE-oxidizing activity occurred in resting cells grown on C5 -C8 n-alkanes. Lower activities occurred in cells grown on longer-chain n-alkanes (C9 -C11 ) and 1°-alcohols (C5 -C10 ). N-octane-grown cells also oxidized tertiary amyl methyl ether (TAME) to tertiary amyl alcohol (TAA), but did not oxidize ethyl tertiary butyl ether (ETBE), TBA or TAA. A 39 kDa polypeptide in whole cell extracts of n-octane-grown cells strongly cross-reacted with an anti-AlkB polyclonal antiserum in an SDS-PAGE/immunoblot. This polypeptide was absent or less abundant in cells grown on dextrose, dextrose plus dicyclopropylketone or 1-octanol. N-octane-grown cells of Pseudomonas aeruginosa strains KSLA-473 and ATCC 17423 oxidized MTBE and TAME but not ETBE. N-hexadecane-grown cells of these strains and strain PAO1 did not oxidize any of the oxygenates tested. Our results indicate ether oxygenate-degrading activity in alkane-utilizing pseudomonads is consistently observed with close homologues of the GPo1 non-haem-iron alkane hydroxylases but is otherwise not a consistent catalytic feature of these diverse enzymes.

Effects of gasoline components on MTBE and TBA cometabolism by Mycobacterium austroafricanum JOB5.

In this study we have examined the effects of individual gasoline hydrocarbons (C(5-10,12,14) n-alkanes, C(5-8) isoalkanes, alicyclics [cyclopentane and methylcyclopentane] and BTEX compounds [benzene, toluene, ethylbenzene, m-, o-, and p-xylene]) on cometabolism of methyl tertiary butyl ether (MTBE) and tertiary butyl alcohol (TBA) by Mycobacterium austroafricanum JOB5. All of the alkanes tested supported growth and both MTBE and TBA oxidation. Growth on C(5-8) n-alkanes and isoalkanes was inhibited by acetylene whereas growth on longer chain n-alkanes was largely unaffected by this gas. However, oxidation of both MTBE and TBA by resting cells was consistently inhibited by acetylene, irrespective of the alkane used as growth-supporting substrate. A model involving two separate but co-expressed alkane-oxidizing enzyme systems is proposed to account for these observations. Cyclopentane, methylcyclopentane, benzene and ethylbenzene did not support growth but these compounds all inhibited MTBE and TBA oxidation by alkane-grown cells. In the case of benzene, the inhibition was shown to be due to competitive interactions with both MTBE and TBA. Several aromatic compounds (p-xylene > toluene > m-xylene) did support growth and cells previously grown on these substrates also oxidized MTBE and TBA. Low concentrations of toluene (<10 microM) stimulated MTBE and TBA oxidation by alkane-grown cells whereas higher concentrations were inhibitory. The effects of acetylene suggest strain JOB5 also has two distinct toluene-oxidizing activities. These results have been discussed in terms of their impact on our understanding of MTBE and TBA cometabolism and the enzymes involved in these processes in mycobacteria and other bacteria.

Isotopic fractionation of methyl tert-butyl ether suggests different initial reaction mechanisms during aerobic biodegradation.

Carbon isotopic enrichment factors (epsilonC) measured during cometabolic biodegradation of methyl tert-butyl ether (MTBE), ethyl tert-butyl ether (ETBE), and tert-amyl methyl ether (TAME) by Pseudonocardia tetrahydrofuranoxydans strain K1 were -2.3 +/- 0.2 per thousand, -1.7 +/- 0.2 per thousand, and -1.7 +/- 0.3 per thousand, respectively. The measured carbon apparent kinetic isotope effect was 1.01 for all compounds, consistent with the expected kinetic isotope effects for both oxidation of the methoxy (or ethoxy) group and enzymatic SN1 biodegradation mechanisms. Significantly, delta13C measurements of the tert-butyl alcohol and tert-amyl alcohol products indicated that the tert-butyl and tert-amyl groups do not participate in the reaction and confirmed that ether biodegradation by strain K1 involves oxidation of the methoxy (or ethoxy) group. Measured hydrogen isotopic enrichment factors (epsilonH) were -100 +/- 10 per thousand, -73 +/- 7 per thousand, and -72 +/- 20 per thousand for MTBE, ETBE, and TAME respectively. Previous results reported for aerobic biodegradation of MTBE by Methylibium petroleiphilum PM1 and Methylibium R8 showed smaller epsilonH values (-35 per thousand and -42 per thousand, respectively). Plots of Delta2H/Delta13C show different slopes for strain K1 compared with strains PM1 and R8, suggesting that different mechanisms are utilized by K1 and PM1/R8 during aerobic MTBE biodegradation.

Pathway, inhibition and regulation of methyl tertiary butyl ether oxidation in a filamentous fungus, Graphium sp.

The filamentous fungus Graphium sp. (ATCC 58400) co-metabolically oxidizes the gasoline oxygenate methyl tertiary butyl ether (MTBE) after growth on gaseous n-alkanes. In this study, the enzymology and regulation of MTBE oxidation by propane-grown mycelia of Graphium sp. were further investigated and defined. The trends observed during MTBE oxidation closely resembled those described for propane-grown cells of the bacterium Mycobacterium vaccae JOB5. Propane-grown mycelia initially oxidized the majority ( approximately 95%) of MTBE to tertiary butyl formate (TBF), and this ester was biotically hydrolyzed to tertiary butyl alcohol (TBA). However, unlike M. vaccae JOB5, our results collectively suggest that propane-grown mycelia only have a limited capacity to degrade TBA. None of the products of MTBE exerted a physiologically relevant regulatory effect on the rate of MTBE or propane oxidation, and no significant effect of TBA was observed on the rate of TBF hydrolysis. Together, these results suggest that the regulatory effects of MTBE oxidation intermediates proposed for MTBE-degrading organisms such as Mycobacterium austroafricanum are not universally relevant mechanisms for MTBE-degrading organisms. The results of this study are discussed in terms of their impact on our understanding of the diversity of aerobic MTBE-degrading organisms and pathways and enzymes involved in these processes.

Propane and n-butane oxidation by Pseudomonas putida GPo1.

Propane and n-butane inhibit methyl tertiary butyl ether oxidation by n-alkane-grown Pseudomonas putida GPo1. Here we demonstrate that these gases are oxidized by this strain and support cell growth. Both gases induced alkane hydroxylase activity and appear to be oxidized by the same enzyme system used for the oxidation of n-octane.

Oxidation of methyl tert-butyl ether by alkane hydroxylase in dicyclopropylketone-induced and n-octane-grown Pseudomonas putida GPo1.

The alkane hydroxylase enzyme system in Pseudomonas putida GPo1 has previously been reported to be unreactive toward the gasoline oxygenate methyl tert-butyl ether (MTBE). We have reexamined this finding by using cells of strain GPo1 grown in rich medium containing dicyclopropylketone (DCPK), a potent gratuitous inducer of alkane hydroxylase activity. Cells grown with DCPK oxidized MTBE and generated stoichiometric quantities of tert-butyl alcohol (TBA). Cells grown in the presence of DCPK also oxidized tert-amyl methyl ether but did not appear to oxidize either TBA, ethyl tert-butyl ether, or tert-amyl alcohol. Evidence linking MTBE oxidation to alkane hydroxylase activity was obtained through several approaches. First, no TBA production from MTBE was observed with cells of strain GPo1 grown on rich medium without DCPK. Second, no TBA production from MTBE was observed in DCPK-treated cells of P. putida GPo12, a strain that lacks the alkane-hydroxylase-encoding OCT plasmid. Third, all n-alkanes that support the growth of strain GPo1 inhibited MTBE oxidation by DCPK-treated cells. Fourth, two non-growth-supporting n-alkanes (propane and n-butane) inhibited MTBE oxidation in a saturable, concentration-dependent process. Fifth, 1,7-octadiyne, a putative mechanism-based inactivator of alkane hydroxylase, fully inhibited TBA production from MTBE. Sixth, MTBE-oxidizing activity was also observed in n-octane-grown cells. Kinetic studies with strain GPo1 grown on n-octane or rich medium with DCPK suggest that MTBE-oxidizing activity may have previously gone undetected in n-octane-grown cells because of the unusually high K(s) value (20 to 40 mM) for MTBE.

Induction of methyl tertiary butyl ether (MTBE)-oxidizing activity in Mycobacterium vaccae JOB5 by MTBE.

Alkane-grown cells of Mycobacterium vaccae JOB5 cometabolically degrade the gasoline oxygenate methyl tertiary butyl ether (MTBE) through the activities of an alkane-inducible monooxygenase and other enzymes in the alkane oxidation pathway. In this study we examined the effects of MTBE on the MTBE-oxidizing activity of M. vaccae JOB5 grown on diverse nonalkane substrates. Carbon-limited cultures were grown on glycerol, lactate, several sugars, and tricarboxylic acid cycle intermediates, both in the presence and absence of MTBE. In all MTBE-containing cultures, MTBE consumption occurred and tertiary butyl alcohol (TBA) and tertiary butyl formate accumulated in the culture medium. Acetylene, a specific inactivator of alkane- and MTBE-oxidizing activities, fully inhibited MTBE consumption and product accumulation but had no other apparent effects on culture growth. The MTBE-dependent stimulation of MTBE-oxidizing activity in fructose- and glycerol-grown cells was saturable with respect to MTBE concentration (50% saturation level = 2.4 to 2.75 mM), and the onset of MTBE oxidation in glycerol-grown cells was inhibited by both rifampin and chloramphenicol. Other oxygenates (TBA and tertiary amyl methyl ether) also induced the enzyme activity required for their own degradation in glycerol-grown cells. Presence of MTBE also promoted MTBE oxidation in cells grown on organic acids, compounds that are often found in anaerobic, gasoline-contaminated environments. Experiments with acid-grown cells suggested induction of MTBE-oxidizing activity by MTBE is subject to catabolite repression. The results of this study are discussed in terms of their potential implications towards our understanding of the role of cometabolism in MTBE and TBA biodegradation in gasoline-contaminated environments.

Cometabolism of methyl tertiary butyl ether and gaseous n-alkanes by Pseudomonas mendocina KR-1 grown on C5 to C8 n-alkanes.

Pseudomonas mendocina KR-1 grew well on toluene, n-alkanes (C5 to C8), and 1 degrees alcohols (C2 to C8) but not on other aromatics, gaseous n-alkanes (C1 to C4), isoalkanes (C4 to C6), 2 degrees alcohols (C3 to C8), methyl tertiary butyl ether (MTBE), or tertiary butyl alcohol (TBA). Cells grown under carbon-limited conditions on n-alkanes in the presence of MTBE (42 micromoles) oxidized up to 94% of the added MTBE to TBA. Less than 3% of the added MTBE was oxidized to TBA when cells were grown on either 1 degrees alcohols, toluene, or dextrose in the presence of MTBE. Concentrated n-pentane-grown cells oxidized MTBE to TBA without a lag phase and without generating tertiary butyl formate (TBF) as an intermediate. Neither TBF nor TBA was consumed by n-pentane-grown cells, while formaldehyde, the expected C1 product of MTBE dealkylation, was rapidly consumed. Similar Ks values for MTBE were observed for cells grown on C5 to C8 n-alkanes (12.95 +/- 2.04 mM), suggesting that the same enzyme oxidizes MTBE in cells grown on each n-alkane. All growth-supporting n-alkanes (C5 to C8) inhibited MTBE oxidation by resting n-pentane-grown cells. Propane (Ki = 53 micromoles) and n-butane (Ki = 16 micromoles) also inhibited MTBE oxidation, and both gases were also consumed by cells during growth on n-pentane. Cultures grown on C5 to C8 n-alkanes also exhibited up to twofold-higher levels of growth in the presence of propane or n-butane, whereas no growth stimulation was observed with methane, ethane, MTBE, TBA, or formaldehyde. The results are discussed in terms of their impacts on our understanding of MTBE biodegradation and cometabolism.

Acetylene inhibition of trichloroethene and vinyl chloride reductive dechlorination.

Kinetic studies reported here have shown that acetylene is a potent reversible inhibitor of reductive dehalogenation of trichloroethene (TCE) and vinyl chloride (VC) by a mixed dehalogenating anaerobic culture. The mixed culture was enriched from a contaminated site in Corvallis, OR, and exhibited methanogenic, acetogenic, and reductive dehalogenation activities. The H2-fed culture transformed TCE to ethene via cis-dichloroethene (c-DCE) and VC as intermediates. Batch kinetic studies showed acetylene reversibly inhibited reduction of both TCE and VC, and the levels of inhibition were strongly dependent on acetylene concentrations in both cases. Acetylene concentrations of 192 and 12 microM, respectively, were required to achieve 90% inhibition in rates of TCE and VC transformation at an aqueous concentration of 400 microM. Acetylene also inhibited methane production (90% inhibition at 48 microM) but did not inhibit H2-dependent acetate production. Mass balances conducted during the studies of VC inhibition showed that acetogenesis, VC transformation to ethene, and methane production were responsible for 52%, 47%, and 1% of the H2 consumption, respectively. The results indicate that halorespiration is the dominant process responsible for VC and TCE transformation and that dehalorespiring organisms are the target of acetylene inhibition. Acetylene has potential use as a reversible inhibitor to probe the biological activities of reductive dechlorination and methanogenesis. It can be added to inhibit reactions and then removed to permit reactions to proceed. Thus, it can be a powerful tool for investigating intrinsic and enhanced anaerobic remediation of chloroethenes at contaminated sites. The results also suggest that acetylene produced abiotically by reactions of chlorinated ethenes with zero-valent iron could inhibit the biological transformation of VC to ethene.

Characterization of the initial reactions during the cometabolic oxidation of methyl tert-butyl ether by propane-grown Mycobacterium vaccae JOB5.

The initial reactions in the cometabolic oxidation of the gasoline oxygenate, methyl tert-butyl ether (MTBE), by Mycobacterium vaccae JOB5 have been characterized. Two products, tert-butyl formate (TBF) and tert-butyl alcohol (TBA), rapidly accumulated extracellularly when propane-grown cells were incubated with MTBE. Lower rates of TBF and TBA production from MTBE were also observed with cells grown on 1- or 2-propanol, while neither product was generated from MTBE by cells grown on casein-yeast extract-dextrose broth. Kinetic studies with propane-grown cells demonstrated that TBF is the dominant (> or = 80%) initial product of MTBE oxidation and that TBA accumulates from further biotic and abiotic hydrolysis of TBF. Our results suggest that the biotic hydrolysis of TBF is catalyzed by a heat-stable esterase with activity toward several other tert-butyl esters. Propane-grown cells also oxidized TBA, but no further oxidation products were detected. Like the oxidation of MTBE, TBA oxidation was fully inhibited by acetylene, an inactivator of short-chain alkane monooxygenase in M. vaccae JOB5. Oxidation of both MTBE and TBA was also inhibited by propane (K(i) = 3.3 to 4.4 microM). Values for K(s) of 1.36 and 1.18 mM and for V(max) of 24.4 and 10.4 nmol min(-1) mg of protein(-1) were derived for MTBE and TBA, respectively. We conclude that the initial steps in the pathway of MTBE oxidation by M. vaccae JOB5 involve two reactions catalyzed by the same monooxygenase (MTBE and TBA oxidation) that are temporally separated by an esterase-catalyzed hydrolysis of TBF to TBA. These results that suggest the initial reactions in MTBE oxidation by M. vaccae JOB5 are the same as those that we have previously characterized in gaseous alkane-utilizing fungi.

Cooxidation of naphthalene and other polycyclic aromatic hydrocarbons by the nitrifying bacterium, Nitrosomonas europaea.

The soil nitrifying bacterium Nitrosomonas europaea has shown the ability to transform cometabolically naphthalene as well as other 2- and 3-ringed polycyclic aromatic hydrocarbons (PAHs) to more oxidized products. All of the observed enzymatic reactions were inhibited by acetylene, a selective inhibitor of ammonia monooxygenase (AMO). A strong inhibitory effect of naphthalene on ammonia oxidation by N. europaea was observed. Naphthalene was readily oxidized by N. europaea and 2-naphthol was detected as a major product (85%) of naphthalene oxidation. The maximum naphthol production rate was 1.65 nmole/mg protein-min in the presence of 240 microM naphthalene and 10 mM NH4+. Our results demonstrate that the oxidation between ammonia and naphthalene showed a partial competitive inhibition. The relative ratio of naphthalene and ammonia oxidation, depending on naphthalene concentrations, demonstrated that the naphthalene was oxidized 2200-fold slower than ammonia at lower concentration of naphthalene (15 microM) whereas naphthalene was oxidized only 100-fold slower than ammonia oxidation. NH4(+)- and N2H4-dependent O2 uptake measurement demonstrated irreversible inhibitory effects of the naphthalene and subsequent oxidation products on AMO and HAO activity.

Biodegradation of Halogenated Hydrocarbon Fumigants by Nitrifying Bacteria.

Three species of nitrifying bacteria were tested for the ability to degrade the halocarbon fumigants methyl bromide, 1,2-dichloropropane, and 1,2-dibromo-3-chloropropane. The soil nitrifiers Nitrosomonas europaea and Nitrosolobus multiformis degraded all three fumigants, while the marine nitrifier Nitrosococcus oceanus degraded only methyl bromide under the conditions tested. Inhibition of biodegradation by allylthiourea and acetylene, specific inhibitors of ammonia monooxygenase, suggests that ammonia monooxygenase is the enzyme which catalyzes fumigant degradation.