Metabolism of L-beta-lysine by a Pseudomonas. Purification and properties of a deacetylase-thiolesterase utilizing 4-acetamidobutyryl CoA and related compounds

A deacetylase-thiolesterase that cleaves both the amide and thiolester bonds of 4-acetamidobutyryl CoA has been highly purified from extracts of Pseudomonas B4 grown in a medium containing L-beta-lysine (3,6-diaminohexanoate) as the main energy source. The enzyme has a molecular weight of about 275,000 and contains 8 apparently identical subunits of 36,500 daltons. Products of 4-acetamidobutyryl CoA degradation are stoichiometric amounts of CoASH and acetate, variable amounts of 4-aminobutyrate and its lactam, 2-pyrrolidinone, and a little 4-acetamidobutyrate. The relative yields of 4-aminobutyrate and 2-pyrrolidinone are determined by the enzyme level. At high enzyme levels the 4-aminobutyrate/pyrrolidinone ratio is about 2, whereas at low enzyme levels only pyrrolidinone is formed. Under the latter conditions, 4-aminobutyryl CoA accumulates transiently and is converted nonenzymatically to pyrrolidinone and CoASH. Since the enzyme does not form 4-aminobutyrate from synthetic or enzymatically formed 4-aminobutyryl CoA, we conclude that a 4-aminobutyryl CoA-enzyme complex is the actual precursor of 4-aminobutyrate, whereas free 4-aminobutyryl CoA is the precursor of pyrrolidinone. Several analogs of 4-acetamidobutyryl CoA containing different amino acid or amide moieties, and several simple acyl CoA compounds are utilized by the enzyme; 4-propionamidobutyryl CoA and 5-acetamidovaleryl CoA are most readily decomposed. Acetyl CoA is a very poor substrate. 3-Acetamidopropionyl CoA is first converted to acetate and beta-alanyl CoA and the latter compound is slowly hydrolyzed to beta-alanine and CoASH. Little deacetylase-thiolesterase is formed by bacteria grown in absence of beta-lysine, but another thiolesterase, lacking deacetylase activity, is produced. The deacetylase-thiolesterase catalyzes an essential step in the aerobic degradation of L-beta-lysine.     

Enzymes involved in 3,5-diaminohexanoate degradation by Brevibacterium sp.

Cell-free extracts of Brevibacterium sp. L5 grown on DL-erythro-3,5-diaminohexanoate were found to contain a 3-keto-5-aminohexanoate cleavage enzyme that converts 3-keto-5-aminohexanoate and acetyl-coenzyme A (CokA) to 3-aminobutyryl-CoA and acetoacetate and a deaminase that coverts L-3-aminobutyryl-CoA to crotonyl-CoA. The cleavage enzyme has been purified extensively, and some of its properties have been determined for comparison with the 3-keto-6-acetamido-hexanoate cleavage enzyme of Pseudomonas sp. B4. The deaminase has been partially purified and characterized. Both the cleavage enzyme and the deaminase are induced by growth on 3,5-diaminohexanoate. The presence of these and other accessory enzymes in Brevibacterium sp. extracts accounts for the results of earlier tracer experiments which showed that C-1 and C-2 of 3-keto-5-aminohexanoate are converted mainly to acetoacetate and acetate, whereas C-3 to C-6 are converted mainly to 3-hydroxybutyrate or its coenzyme A thiolester. The enzymes observed in extracts of Brevibacterium sp. can account for the conversion of 3,5-diaminohexanoate to acetyl-CoA.     

Variables associated with peripartum traits in dairy cows. III. Effect of diets and disorders on certain blood traits

BLood samples were collected from 89 Holstein cows on days 220 and 250 of gestation, within 24 hr prepartum and postpartum and on day 30 postpartum. Balanced diets which contained either chopped hay (29 cows), hay crop silage (HCS; 30 cows) or corn silage (CS; 30 cows) were fed from day 220 of gestation to day 30 postpartum. The purpose was to determine if variations in certain blood traits were indicative of peripartum and postpartum disorders. The blood traits evaluated were concentrations of plasma total protein, whole blood hemoglobin, packed cell volume, and white blood cells, and serum glutamic oxalacetic transaminase (SGOT), glucose, urea nitrogen, calcium, inorganic phosphorus, magnesium, potassium and sodium. No blood trait was useful to predict a disorder prior to its visual signs with one possible exception. Serum glucose and calcium were lower and SGOT and magnesium were higher peripartum which was prior to death of three cows from fat cow syndrome.     

Assessment of genetic stability on in vitro and ex vitro plants of Ficus carica var. black jack using ISSR and DAMD markers

Molecular Biology Reports - Clonal propagation is one of the attributes of plant tissue culture. Therefore, analysis of genetic stability among the in vitro cultured plants is a crucial step. It...     

Phagocytosis of degenerating myelin in transected feline optic nerve: an immunohistochemical study

The phagocytic activity of neuroglial cells in adult feline degenerating optic nerve was investigated by immunocytochemistry at both light and electron microscopy levels. Degeneration was initiated by unilateral eye enucleation and the segment distal to the transection showing true Wallerian degeneration was examined. Following enucleation, twelve adult domestic cats were examined over a period of seven to 215 days. All cases showed slow clearance of myelin debris and absence of proliferating monocytes throughout the post-enucleation period. All phagocytic cells present were neuroglial cells, and many of these cells expressed oligodendroglial antigens. These findings demonstrate the persistence of an active population of oligodendrocytes that might play an additional functional role during Wallerian degeneration of feline optic nerve.     

Properties and Expression of Na+/K+-ATPase α-Subunit Isoforms in the Brain of the Swamp Eel,Monopterus albus,Which Has Unusually High Brain Ammonia Tolerance

The swamp eel, Monopterus albus, can survive in high concentrations of ammonia (>75 mmol l⁻¹) and accumulate ammonia to high concentrations in its brain (∼4.5 µmol g⁻¹). Na⁺/K⁺-ATPase (Nka) is an essential transporter in brain cells, and since NH₄ ⁺ can substitute for K⁺ to activate Nka, we hypothesized that the brain of M. albus expressed multiple forms of Nka α-subunits, some of which might have high K⁺ specificity. Thus, this study aimed to clone and sequence the nka α-subunits from the brain of M. albus, and to determine the effects of ammonia exposure on their mRNA expression and overall protein abundance. The effectiveness of NH₄ ⁺ to activate brain Nka from M. albus and Mus musculus was also examined by comparing their Na⁺/K⁺-ATPase and Na⁺/NH₄ ⁺-ATPase activities over a range of K⁺/NH₄ ⁺ concentrations. The full length cDNA coding sequences of three nkaα (nkaα1, nkaα3a and nkaα3b) were identified in the brain of M. albus, but nkaα2 expression was undetectable. Exposure to 50 mmol l⁻¹ NH₄Cl for 1 day or 6 days resulted in significant decreases in the mRNA expression of nkaα1, nkaα3a and nkaα3b. The overall Nka protein abundance also decreased significantly after 6 days of ammonia exposure. For M. albus, brain Na⁺/NH₄ ⁺-ATPase activities were significantly lower than the Na⁺/K⁺-ATPase activities assayed at various NH₄ ⁺/K⁺ concentrations. Furthermore, the effectiveness of NH₄ ⁺ to activate Nka from the brain of M. albus was significantly lower than that from the brain of M. musculus, which is ammonia-sensitive. Hence, the (1) lack of nkaα2 expression, (2) high K⁺ specificity of K⁺ binding sites of Nkaα1, Nkaα3a and Nkaα3b, and (3) down-regulation of mRNA expression of all three nkaα isoforms and the overall Nka protein abundance in response to ammonia exposure might be some of the contributing factors to the high brain ammonia tolerance in M. albus.     

The fluted giant clam (Tridacna squamosa) increases nitrate absorption and upregulates the expression of a homolog of SIALIN (H+:2NO3− cotransporter) in the ctenidium during light exposure

Coral Reefs - Giant clams flourish in nutrient-poor waters of tropical Indo-Pacific because they live in symbiosis with extracellular dinoflagellates (zooxanthellae) and receive photosynthates from...     

Diabetes-Related Distress,Depression and Distress-Depression among Adults with Type 2 Diabetes Mellitus in Malaysia

Type 2 diabetes mellitus (T2DM) brings about an increasing psychosocial problem in adult patients. Prevalence data on and associated factors of diabetes related distress (DRD) and depression have been lacking in Asia. This study aimed to examine the prevalence of DRD and depression, and their associated factors in Asian adult T2DM patients. This study was conducted in three public health clinics measuring DRD (Diabetes Distress Scale, DDS), and depression (Patient Health Questionnaire, PHQ). Patients who were at least 30 years of age, had T2DM for more than one year, with regular follow-up and recent laboratory results (< 3 months) were consecutively recruited. Associations between DRD, depression and the combination DRD-depression with demographic and clinical characteristics were analysed using generalized linear models. From 752 invited people, 700 participated (mean age 56.9 years, 52.8% female, 52.9% Malay, 79.1% married). Prevalence of DRD and depression were 49.2% and 41.7%, respectively. Distress and depression were correlated, spearman’s r = 0.50. Patients with higher DRD were younger (OR 0.995, 95% CI 0.996 to 0.991), Chinese (OR 1.2, 95% CI 1.04 to 1.29), attending Dengkil health clinic (OR 1.1, 95% CI 1.00 to 1.22) and had higher scores on the PHQ (OR 1.1, 95% CI 1.04 to 1.06). Depression was less likely in the unmarried compared to divorced/separately living and those attending Dengkil health clinic, but more likely in patients with microvascular complications (OR 1.4, 95% CI 1.06 to 1.73) and higher DDS (OR 1.03, 95% CI 1.02 to 1.03). For the combination of DRD and depression, unemployment (OR 4.7, 95% CI 1.02 to 21.20) had positive association, whereas those under medical care at the Salak health clinics (OR 0.28, 95% CI 0.12 to 0.63), and those with a blood pressure > 130/80 mmHg (OR 0.53, 95% CI 0.32 to 0.89) were less likely to experience both DRD and depression. DRD and depression were common and correlated in Asian adults with T2DM at primary care level. Socio-demographic more than clinical characteristics were related to DRD and depression.