Geochemistry of Liquid Hydrocarbons and Natural Gases Combined with 1D Basin Modeling of the Oligocene Shale Source Rock System in the Offshore Nile Delta, Egypt.

The current study aims to integrate the geochemical characteristics of the Oligocene shale source rock system, oil, condensate, and natural gas samples in the Oligocene sandstone reservoirs from three exploration wells located in the offshore Nile Delta, East Mediterranean Sea, using organic geochemistry and a 1D basin modeling scheme. The Tineh shales exhibit total organic carbon values ranging between 0.90 and 1.89 wt %, along with hydrogen index values in the range of 54-240 mg hydrocarbon/g rock. The geochemical characterization suggests that the shale intervals of the Oligocene Tineh Formation contain type II-III and type III kerogens and, thereby, could be regarded as promising oil- and gas-prone source rocks with high contributions of gas generation potential. The study also reconstructs the 1D thermal and burial history models, showing that the Oligocene Tineh source rock system is in the main oil and wet gas generation phases from the late Miocene to the present time. The simulated basin models reveal the transformation (TR) of 10-50% kerogen to oil during the late Miocene-early Pliocene period and that the Oligocene Tineh source rock system has larger oil generation and expulsion competency, with a TR value of up to 65% during the early Pliocene-Pleistocene time period. The thermogenic gas was also formed during this time and continued to the present day. This study also investigated the presence of oil and condensate in the Oligocene sandstone reservoir samples and revealed that they were generated from mature source rock, ranging from moderately to highly mature stages. This source rock unit was deposited in fluvial to fluvial-deltaic environments under oxic mixed organic conditions and accumulated during the Tertiary time, as evidenced by the presence of the oleanane biomarker dating indicator. The molecular and isotope compositions of natural gases revealed that most of the natural gases in the Oligocene sandstone reservoir are mainly thermogenic methane gases that were generated from mainly mixed organic matter. The thermogenic methane gases were formed mainly from secondary cracking of oil and gas, with small contributions of primary kerogen cracking. The properties of natural gases together with oil and condensate in the Oligocene reservoir rocks suggest that most of the thermogenic methane gases and associated liquid hydrocarbons are derived primarily from the Oligocene shale source rock system and formed by primary kerogen cracking and secondary oil and oil/gas cracking in different thermal maturity stages. Therefore, the Oligocene Tineh Formation can be regarded as self-source generation and self-reservoir rock; hence, an intensive oil exploration and production program can be recommended whenever the Tineh source rock system is is well developed and deeply buried.

Collaborative Impact of Compost and Beneficial Rhizobacteria on Soil Properties, Physiological Attributes, and Productivity of Wheat Subjected to Deficit Irrigation in Salt Affected Soil.

Plant growth and crop productivity under unfavorable environmental challenges require a unique strategy to scavenge the severely negative impacts of these challenges such as soil salinity and water stress. Compost and plant growth-promoting rhizobacteria (PGPR) have many beneficial impacts, particularly in plants exposed to different types of stress. Therefore, a field experiment during two successive seasons was conducted to investigate the impact of compost and PGPR either separately or in a combination on exchangeable sodium percentage (ESP), soil enzymes (urease and dehydrogenase), wheat physiology, antioxidant defense system, growth, and productivity under deficient irrigation and soil salinity conditions. Our findings showed that exposure of wheat plants to deficit irrigation in salt-affected soil inhibited wheat growth and development, and eventually reduced crop productivity. However, these injurious impacts were diminished after soil amendment using the combined application of compost and PGPR. This combined application enhanced soil urease and dehydrogenase, ion selectivity, chlorophylls, carotenoids, stomatal conductance, and the relative water content (RWC) whilst reducing ESP, proline content, which eventually increased the yield-related traits of wheat plants under deficient irrigation conditions. Moreover, the coupled application of compost and PGPR reduced the uptake of Na and resulted in an increment in superoxide dismutase (SOD), catalase (CAT), and peroxidase (POX) activities that lessened oxidative damage and improved the nutrient uptake (N, P, and K) of deficiently irrigated wheat plants under soil salinity. It was concluded that to protect wheat plants from environmental stressors, such as water stress and soil salinity, co-application of compost with PGPR was found to be effective.

End-to-end anal sphincter repair in treatment of post-traumatic fecal incontinence

The present report describes the end-to-end technique of anal sphincter repair in a 36-year-old female patient with post-vaginal delivery fecal incontinence (FI). The patient had a history of two vaginal deliveries and the symptoms of FI were observed after the second delivery. On assessment of the severity of FI using the Wexner incontinence score, the patient had a score of 12. Endoanal ultrasonography revealed an anterior defect of the external anal sphincter extending from 11 to 3 o'clock. The patient had no previous anal surgery and did not have any medical comorbidities. The operation time was 45minutes. No intraoperative complications were recorded. At 12 months of follow-up, the patient showed significant improvement in the continence state, with her Wexner score dropping to 4. No postoperative complications were recorded. We can conclude that end-to-end anal sphincter repair is a technically feasible operation that confers satisfactory improvement in the continence state without imposing much tension on the site of sphincter repair. (AU)

Complete Substitution of Clover Hay and its Impact on Performance of Barki Sheep in Arid and Semi-arid Regions.

BACKGROUND AND OBJECTIVE: Barley straw is a farm by product that may be useful in animal feeding but its use is limited due to its low nutritional value and high fiber content. Therefore, this study aimed to improving the nutritive value of barley straw by bacterial treatment with Condensed Molasses Soluble (CMS) in the ration of Barki lambs. MATERIALS AND METHODS: In Experiment 1, 12 Barki rams, which were divided into 3 equal groups (4 each), were given rations which consisted of 50% CFM+50%, either clover hay (C) or bacterial treated barley straw (T1) or T1+ CMS (T2). In Experiment 2, 10 Barki lambs were divided into 2 groups, one (C) and T2 as descripted in Experiment 1. Data analyzed by using one way ANOVA model. RESULTS: Digestibility of Dry Matter (DM), Organic Matter (OM), Crude Fiber (CF) and Ether Extract (EE) were insignificantly affected among all rations, while Digestible Crude Protein (DCP) recorded a higher value in T2 compared to T1. Changes in Total Digestible Nutrients (TDN) values were insignificant. Values of DCP were higher for T2 compared to C and T1. The Body Weight Gain (BWG) and Feed Conversion Ratio (FCR) of growing lambs didn't statistically differ among the groups. CONCLUSION: Finally, complete replacement of clover hay by treated barley straw plus CMS may be considered good feeding strategies in the feeding of Barki lambs in Arid and semi-arid regions.

Modification of Parks Classification of Cryptoglandular Anal Fistula.

BACKGROUND: The Parks classification has been used for the classification of anal fistula for several years, but it does not allow for risk factors for failure after surgery. OBJECTIVE: This study aimed to describe a modification of the Parks classification of anal fistula and examine its predictive validity in the assessment of the outcome of anal fistula in terms of failure of healing and fecal incontinence. DESIGN: This is a retrospective review of a prospective database. SETTING: This study was conducted in the Colorectal Surgery Unit, Mansoura University Hospitals. PATIENTS: Adult patients with anal fistula who underwent surgery were included. INTERVENTIONS: Five risk factors for failure after fistula surgery were identified from the literature and were examined by multivariate analysis of our patients. Four risk factors proved to be significant independent predictors of failure: secondary extensions, horseshoe fistula, previous fistula surgery, and anterior anal fistula in women. We modified the Parks classification by dividing the transsphincteric type into high and low and by grouping supra- and extrasphincteric anal fistulas into 1 group. The first 3 stages were subdivided according to the absence or presence of predictors of failure. MAIN OUTCOME MEASURES: The primary outcome measured was the validity of the modified Parks classification with regard to the rates of failure and fecal incontinence after surgical treatment of each stage of anal fistula. RESULTS: A total of 665 patients with cryptoglandular anal fistula were included. Failure rates increased from 2.3% (95% CI, 0.9%-4.7%), to 17.4% (95% CI, 10.8%-25.9%), 19.5% (95% CI, 15%-24.6%), and 30.7% (95% CI, 9.1%-61.4%) across the 4 stages. The area under the receiver operating characteristic curve was 0.90 (95% CI, 0.85-0.94) indicating the strong discriminative ability of the final multivariable predictive model. The increase in failure and incontinence rates across the fistula stages was significant. LIMITATIONS: This is a retrospective, single-center study. CONCLUSION: Inclusion of predictors of poor outcome into the modified classification helped differentiate simple and complex fistulas within each stage and between the different stages, which can help in assessment and decision making for anal fistula. See Video Abstract at http://links.lww.com/DCR/B441. MODIFICACIN DE LA CLASIFICACIN DE PARKS DE LA FSTULA ANAL CRIPTOGLANDULAR: ANTECEDENTES:La clasificación de Parks se ha utilizado para la clasificación de la fístula anal durante varios años, sin embargo, no tuvo en cuenta los factores de riesgo de fracaso después de la cirugía.OBJETIVO:Describir una modificación de la clasificación de Parks de fístula anal y examinar su validez predictiva en la evaluación de los resultados de la fístula anal en términos de fracaso de la cicatrización e incontinencia fecal.DISEÑO:Revisión retrospectiva de la base de datos prospectiva.AJUSTE:Unidad de Cirugía Colorrectal, Hospital Universitario de Mansoura.PACIENTES:Pacientes adultos con fístula anal intervenidos quirúgicamente.INTERVENCIONES:Se identificaron cinco factores de riesgo de fracaso después de la cirugía de fístula de la literatura y se examinaron mediante análisis multivariante de nuestros pacientes. Cuatro factores de riesgo demostraron ser importantes predictores independientes de fracaso: extensiones secundarias, fístula en herradura, cirugía de fístula previa y fístula anal anterior en mujeres. Modificamos la clasificación de Parks dividiendo el tipo transesfinteriano en alto y bajo y agrupando la fístula anal supraesfinteriana y extraesfinteriana en un grupo. Las tres primeras etapas se subdividieron según la ausencia o presencia de predictores de fracaso.PRINCIPALES MEDIDAS DE RESULTADO:Validez de la clasificación de Parks modificada con respecto a las tasas de fracaso e incontinencia fecal después del tratamiento quirúrgico de cada etapa de la fístula anal.RESULTADOS:Se incluyeron 665 pacientes con fístula anal criptoglandular. Las tasas de fracaso aumentaron del 2,3% (IC del 95%: 0,9-4,7%), al 17,4% (IC del 95%: 10,8 al 25,9%), 19,5% (IC del 95%: 15-24,6%) y 30,7% (95% IC: 9,1- 61,4%) en las cuatro etapas. El área bajo la curva característica operativa del receptor fue 0,90 (IC del 95%: 0,85-0,94), lo que indica una fuerte capacidad discriminativa del modelo predictivo multivariable final. El aumento en las tasas de fracaso e incontinencia en las etapas de la fístula fue significativo.LIMITACIONES:Estudio retrospectivo, unicéntrico.CONCLUSIÓN:La inclusión de predictores de mal resultado en la clasificación modificada ayudó a diferenciar las fístulas simples y complejas dentro de cada etapa y entre las diferentes etapas, lo que puede ayudar en la evaluación y toma de decisiones para la fístula anal. Consulte Video Resumen en http://links.lww.com/DCR/B441.

Video demonstration of the modified Parks' technique for treatment of trans-sphincteric anal fistula.

BACKGROUND: Trans-sphincteric anal fistula (TAF) is a common type of anal fistula that can be divided into low and high subtypes. Sphincter-sparing surgery is usually used for treatment of high TAF to avoid any compromise of the anal continence. METHODS: A 38-year-old male patient with recurrent Grade III TAF was treated with the modified Parks' technique. This technique can be summarized as three main steps; each addresses a component of the anal fistula. (1) The extra-sphincteric component, which lies outside the external anal sphincter (EAS), is laid open with electrocautery. (2) The intersphincteric component is laid open, starting from the internal opening, dividing the internal anal sphincter, until the EAS fibers were reached. (3) The trans-sphincteric component that traverses the EAS was carefully curetted from the anal and the outer sides with a surgical curette. An important step of the procedure is to close the defect through which the fistula tract passed through the EAS with interrupted sutures. RESULTS: The operation time was 35 min. Complete healing was achieved in 5 weeks with no persistence or recurrence of the anal fistula after a follow-up of 6 months. No affection of the continence state was recorded. CONCLUSION: The modified Parks' technique is a simple and convenient operation for TAF. Further prospective studies are needed to substantiate the efficacy and safety of the procedure.

Horizontal versus vertical plication of the rectovaginal septum in transperineal repair of anterior rectocele: a pilot randomized clinical trial.

AIM: Anterior rectocele is usually an asymptomatic condition in many women, yet it can be associated with obstructed defaecation syndrome (ODS). Transperineal repair of rectocele (TPR) has been followed by variable rates of improvement in ODS. The present pilot randomized clinical trial aimed to evaluate the outcome of TPR with vertical plication (VP) of the rectovaginal septum compared to horizontal plication (HP). METHODS: Adult women with anterior rectocele were recruited to the study and were randomly allocated to one of two equal groups. The first group underwent TPR with VP of the rectovaginal septum and the second group underwent TPR with HP. The main outcome measures were improvement in ODS, recurrence of rectocele, complications and dyspareunia. RESULTS: The trial included 40 female patients with anterior rectocele. There was no significant difference between the two groups regarding the postoperative Wexner score. Complete cure and significant improvement in ODS symptoms were comparable after the two techniques. The reduction in rectocele size after HP was significantly greater than after VP (1.7 vs. 2.6, P < 0.0001). Significant improvement in dyspareunia was recorded after HP (P = 0.001) but not after VP (P = 0.1). There was no significant difference between the two groups with regard to operating time, complications and recurrence. CONCLUSION: VP and HP of the rectovaginal septum in TPR were associated with a comparable improvement in ODS symptoms and similar complication rates. HP was followed by a greater reduction in the rectocele size and greater improvement in dyspareunia than VP.

Effect of harvest season on the nutritional value of bee pollen protein.

Bee pollen is a natural product that has valuable nutritional and medicinal characteristics and has recently garnered increasing attention in the food industry due to its nutritive value. Here, we harvested pollen loads from the Al-Ahsa oasis in eastern Saudi Arabia during spring, summer, autumn, and winter in 2018/2019 to compare the nutritional value of bee pollen protein with the amino acid requirements of honeybees and adult humans. Based on the nutritional value of bee pollen protein, the optimal season for harvesting bee pollen was determined. The composition of the bee pollen showed the highest contents of crude protein, total amino acids, leucine, glutamic acid, valine, isoleucine, threonine, and glycine in samples collected in spring. The highest contents of lysine, phenylalanine, threonine, tryptophan, arginine, tyrosine, and cysteine were observed in samples collected in winter. The highest contents of histidine, methionine, and serine were in samples collected in autumn. Moreover, the highest levels of aspartic acid, proline, and alanine were in samples collected in summer. Leucine, valine, lysine, histidine, threonine, and phenylalanine (except in autumn bee pollen) contents in pollen from all four seasons were above the requirements of honeybees. Leucine, valine, histidine, isoleucine (except in autumn bee pollen), lysine (except in spring and summer bee pollen), and threonine (except in winter and spring bee pollen) in all tested samples were above the requirements of adult humans. In comparison with the minimal amino acid requirements of adult humans and honeybees, the 1st limiting amino acid in bee pollen collected during the different seasons was methionine. Bee pollen collected during spring (March-May) and winter (December-February) can be considered a nutritive food source for adult humans and honeybees.

A randomized controlled trial on irrigation of open appendectomy wound with gentamicin- saline solution versus saline solution for prevention of surgical site infection.

BACKGROUND: Surgical site infection (SSI) is one of the most common complications after abdominal surgery. The present trial examined the efficacy of saline irrigation of open appendectomy wound with or without topical antibiotics in prevention of SSI. METHODS: This was a double-blind randomized trial on patients with acute appendicitis who underwent open appendectomy. Patients were randomly allocated to one of three equal groups; group I had layer-by-layer wound irrigation with gentamicin-saline solution, group II had wound irrigation with saline solution, and group III received no irrigation (Control group). The main outcome measures were the incidence of incisional SSI, surgical site occurrence (SSO), other complications, operation time, postoperative pain, and patients' satisfaction. RESULTS: 205 patients (113 female) of a mean age of 27.9 years were included. The average hospital stay and pain scores were similar in the three groups. Groups I and II had significantly lower rates of incisional SSI (4.3% Vs 2.9%; Vs 17.4%, p = 0.005) and SSO (24.6% Vs 13.4% Vs 43.5%; p = 0.0003) as compared to group III. Groups I and II had comparable rates of SSI and SSO. The three groups had similar rates of wound seroma, hematoma, and dehiscence. Groups I and II had significantly higher satisfaction with the procedure than group III. CONCLUSIONS: Layer-by-layer irrigation of open appendectomy wound decreased the rates of incisional SSI and SSO significantly compared to the no-irrigation group. Adding gentamicin to saline solution was useless to improve the outcome and did not decrease rates of SSI or other complications.

Drained mucosal advancement flap versus rerouting Seton around the internal anal sphincter in treatment of high trans-sphincteric anal fistula: A randomized trial.

BACKGROUND: Several sphincter saving techniques have been described for complex anal fistula (CAF) with variable outcomes. The present trial aimed to compare two techniques for CAF; the drained mucosal flap technique and rerouting Seton around the internal anal sphincter (IAS). METHODS: Adult patients with high trans-sphincteric anal fistula were randomly assigned to one of two groups: group I underwent mucosal advancement flap with drainage Seton rerouted around the external anal sphincter, and group II underwent rerouting Seton around the IAS. The two groups were compared in terms of the incidence of postoperative fecal incontinence (FI), healing of fistula, complications, and changes in anal pressures. RESULTS: 97 patients (80 male) of a mean age of 39.5 years were included. One patient developed FI in group I versus 7 in group II (p = 0.03). Failure of healing occurred in 2 patients in group I and 4 in group II (p = 0.43). In group II, the average time for spontaneous fall of Seton was 14 ± 2.8 days whereas in group I the average time for removal of Seton was 40 ± 14.9 days (p < 0.0001). There were no significant differences between the two groups in complication rate. Postoperatively, the decrease in resting anal pressure was significant in Group II but not group I. CONCLUSION: The drained mucosal flap technique was associated with significantly lower incidence of FI, yet longer operative time and longer time to complete healing compared to rerouting Seton around the IAS. The success rates of both techniques was comparable.

Rotational gluteal flap versus modified Limberg flap in treatment of sacrococcygeal pilonidal disease.

BACKGROUND: Sacrococcygeal pilonidal disease (SPND) is a common surgical condition with a multitude of surgical treatments. The aim of this study was to compare the outcome of modified Limberg flap (MLF) and that of rotational gluteal flap (RGF) in treatment of SPND. MATERIALS AND METHODS: This was a retrospective case-controlled study involving adult patients of both genders who were treated with either MLF or RGF for SPND. The main outcomes of the study were the recurrence of SPND, complications, operation time, hospital stay, time of healing, time to return to daily activities, and cosmetic outcomes. RESULTS: A total of 189 (143 males) patients with a mean age of 23.5 y were included. Ninety-five patients were treated with RGF (cases), and 94 were treated with MLF (controls). Both the groups were matched regarding age, gender, body mass index, and disease severity. The operation time of RGF was longer than that of MLF (P < 0.0001). No significant difference in recurrence was noted between RGF and MLF (3.1% versus 7.4%, P = 0.21). RGF had lower complication rate than MLF (17.9% versus 40.4%; P = 0.001). Time to return to work was similar in both the groups. RGF conferred significantly better cosmetic outcomes than MLF. CONCLUSIONS: Both RGF and MLF achieved comparable, low recurrence rates. Although the operation time of RGF was significantly longer than that of MLF, the RGF had shorter healing time; lower complication rate, particularly seroma formation; and better cosmetic appearance.

Metabolic and process engineering for biodesulfurization in Gram-negative bacteria.

Microbial desulfurization or biodesulfurization (BDS) is an attractive low-cost and environmentally friendly complementary technology to the hydrotreating chemical process based on the potential of certain bacteria to specifically remove sulfur from S-heterocyclic compounds of crude fuels that are recalcitrant to the chemical treatments. The 4S or Dsz sulfur specific pathway for dibenzothiophene (DBT) and alkyl-substituted DBTs, widely used as model S-heterocyclic compounds, has been extensively studied at the physiological, biochemical and genetic levels mainly in Gram-positive bacteria. Nevertheless, several Gram-negative bacteria have been also used in BDS because they are endowed with some properties, e.g., broad metabolic versatility and easy genetic and genomic manipulation, that make them suitable chassis for systems metabolic engineering strategies. A high number of recombinant bacteria, many of which are Pseudomonas strains, have been constructed to overcome the major bottlenecks of the desulfurization process, i.e., expression of the dsz operon, activity of the Dsz enzymes, retro-inhibition of the Dsz pathway, availability of reducing power, uptake-secretion of substrate and intermediates, tolerance to organic solvents and metals, and other host-specific limitations. However, to attain a BDS process with industrial applicability, it is necessary to apply all the knowledge and advances achieved at the genetic and metabolic levels to the process engineering level, i.e., kinetic modelling, scale-up of biphasic systems, enhancing mass transfer rates, biocatalyst separation, etc. The production of high-added value products derived from the organosulfur material present in oil can be regarded also as an economically viable process that has barely begun to be explored.

Genome Sequence of Pseudomonas azelaica Strain Aramco J.

We report here the draft genome sequence of Pseudomonas azelaica strain Aramco J (7.3 Mbp; GC content, 61.9%), one of the few bacteria that can completely mineralize different hydroxybiphenyls, e.g., 2-hydroxybiphenyl, 2,2'-dihydroxybiphenyl, and 3-hydroxybiphenyl. The findings obtained from its genome annotation suggest that this strain becomes a useful biocatalyst for aromatic bioconversions.

Genome Sequence of Pseudomonas azelaica HBP1, Which Catabolizes 2-Hydroxybiphenyl Fungicide.

Pseudomonas azelaica HBP1 (DSM 8897) is one of the few bacteria able to completely mineralize the 2-hydroxybiphenyl biocide. Here, we report the draft genome sequence of this strain (7.4 Mbp; G+C content, 63.5%) and the findings obtained from its genome annotation.

Ameliorative effect of grapefruit juice on amiodarone-induced cytogenetic and testicular damage in albino rats

Objective:To evaluate the ameliorative role of grapefruit juice on the cytogenetic and testicular damage induced by the antiarrythmic drug amiodarone in albino rats. Methods: Animals were divided into four groups. Group I was considered as control. Group II was given grapefruit juice at a dose level of 27 mL/kg body weight. Group III was orally administered amiodarone (18 mg/kg body weight) daily for 5 weeks. Animals were sacrificed after 5 weeks of treatment. Bone marrow was collected from the femurs for analysis of chromosomal aberrations and mitotic indices. Testes were removed and stained with H&E for histological examination. Sperms were collected from epidedymis for detection of sperm head abnormalities. Comet assay was used to detect DNA damage. Results: Amiodarone treatment caused a significant increase in the percentage of chromosomal aberrations, decreased the mitotic index and increased DNA damage. The testis showed many histopathological alterations, inhibition of spermatogenesis and morphometric changes. The number of sperm head abnormalities was increased. Treating animals with amiodarone and grapefruit juice caused a reduction in chromosomal aberrations, mitotic index, DNA damage and testicular alterations caused by amiodarone. Conclusions:The results of this study indicated that grapefruit juice ameliorates the cytotoxicty and testicular alterations induced by amiodarone in albino rats and this is may be due to the potent antioxidant effects of its components.

Functional genomics by NMR spectroscopy. Phenylacetate catabolism in Escherichia coli.

Aerobic metabolism of phenylalanine in most bacteria proceeds via oxidation to phenylacetate. Surprisingly, the further metabolism of phenylacetate has not been elucidated, even in well studied bacteria such as Escherichia coli. The only committed step is the conversion of phenylacetate into phenylacetyl-CoA. The paa operon of E. coli encodes 14 polypeptides involved in the catabolism of phenylacetate. We have found that E. coli K12 mutants with a deletion of the paaF, paaG, paaH, paaJ or paaZ gene are unable to grow with phenylacetate as carbon source. Incubation of a paaG mutant with [U-13C8]phenylacetate yielded ring-1,2-dihydroxy-1,2-dihydrophenylacetyl lactone as shown by NMR spectroscopy. Incubation of the paaF and paaH mutants with phenylacetate yielded delta3-dehydroadipate and 3-hydroxyadipate, respectively. The origin of the carbon atoms of these C6 compounds from the aromatic ring was shown using [ring-13C6]phenylacetate. The paaG and paaZ mutants also converted phenylacetate into ortho-hydroxyphenylacetate, which was previously identified as a dead end product of phenylacetate catabolism. These data, in conjunction with protein sequence data, suggest a novel catabolic pathway via CoA thioesters. According to this, phenylacetyl-CoA is attacked by a ring-oxygenase/reductase (PaaABCDE proteins), generating a hydroxylated and reduced derivative of phenylacetyl-CoA, which is not re-oxidized to a dihydroxylated aromatic intermediate, as in other known aromatic pathways. Rather, it is proposed that this nonaromatic intermediate CoA ester is further metabolized in a complex reaction sequence comprising enoyl-CoA isomerization/hydration, nonoxygenolytic ring opening, and dehydrogenation catalyzed by the PaaG and PaaZ proteins. The subsequent beta-oxidation-type degradation of the resulting CoA dicarboxylate via beta-ketoadipyl-CoA to succinyl-CoA and acetyl-CoA appears to be catalyzed by the PaaJ, PaaF and PaaH proteins.

Biochemical and molecular characterization of phenylacetate-coenzyme A ligase, an enzyme catalyzing the first step in aerobic metabolism of phenylacetic acid in Azoarcus evansii.

Phenylacetate-coenzyme A ligase (PA-CoA ligase; AMP forming, EC 6.2. 1.30), the enzyme catalyzing the first step in the aerobic degradation of phenylacetate (PA) in Azoarcus evansii, has been purified and characterized. The gene (paaK) coding for this enzyme was cloned and sequenced. The enzyme catalyzes the reaction of PA with CoA and MgATP to yield phenylacetyl-CoA (PACoA) plus AMP plus PPi. The enzyme was specifically induced after aerobic growth in a chemically defined medium containing PA or phenylalanine (Phe) as the sole carbon source. Growth with 4-hydroxyphenylacetate, benzoate, adipate, or acetate did not induce the synthesis of this enzyme. This enzymatic activity was detected very early in the exponential phase of growth, and a maximal specific activity of 76 nmol min(-1) mg of cell protein(-1) was measured. After 117-fold purification to homogeneity, a specific activity of 48 micromol min(-1) mg of protein(-1) was achieved with a turnover number (catalytic constant) of 40 s(-1). The protein is a monomer of 52 kDa and shows high specificity towards PA; other aromatic or aliphatic acids were not used as substrates. The apparent K(m) values for PA, ATP, and CoA were 14, 60, and 45 microM, respectively. The PA-CoA ligase has an optimum pH of 8 to 8.5 and a pI of 6.3. The enzyme is labile and requires the presence of glycerol for stabilization. The N-terminal amino acid sequence of the purified protein showed no homology with other reported PA-CoA ligases. The gene encoding this enzyme is 1, 320 bp long and codes for a protein of 48.75 kDa (440 amino acids) which shows high similarity with other reported PA-CoA ligases. An amino acid consensus for an AMP binding motif (VX2SSGTTGXP) was identified. The biochemical and molecular characteristics of this enzyme are quite different from those of the isoenzyme catalyzing the same reaction under anaerobic conditions in the same bacterium.