Insights into bacterial associations catabolizing atrazine by culture-dependent and molecular approaches

With the specific selection pressures of four atrazine concentrations (10–33 mg l^–1) and two pH values (5.5 and 7.5), eight atrazine-catabolizing microbial associations were enriched and isolated from pesticide-contaminated South African loamy soil. Community-level physiological profiling of Environmental Biolog analysis data identified species complement differences in response to both pH and atrazine concentration and these were confirmed by polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE). These differences were not detected by conventional plate cultures and light and scanning electron microscopy. To probe atrazine catabolism under a range of environmental conditions, the two (pH 5.5, KRA02; pH 7.5, KRA06) associations catabolizing 30 mg atrazine l^–1 were combined (KRA30). The highest specific growth rate was recorded at pH 4, while at pH 8 little growth resulted. With pH 4-poised cultures, the specific growth rates at 15 and 20 °C were comparable but more than doubled for the next 10° increment. These differences reflected species complement changes. Direct comparison of KRA30 with a reference strain, Pseudomonassp. strain ADP, identified comparable specific growth and atrazine catabolic rates. To probe catabolism further, nitrogen-limited batch cultures were made in the presence of supplemental carbon (citrate) but the catabolic rate did not change. The results are discussed with reference to in situ bioaugmentation remediation programmes.     

Mice that lack matrix metalloproteinase-9 display delayed wound healing associated with delayed reepithelization and disordered collagen fibrillogenesis

Matrix metalloproteinase- (MMP-9) is involved in processes that occur during cutaneous wound healing such as inflammation, matrix remodeling, and epithelialization, To investigate its role in healing, full thickness skin wounds were made in the dorsal region of MMP-9-null and control mice and harvested up to 14 days post wounding. Gross examination and histological and immunohistochemical analysis indicated delayed healing in MMP-9-null mice. Specifically, MMP-9-null wounds displayed compromised reepithelialization and reduced clearance of fibrin clots. In addition, they exhibited abnormal matrix deposition, as evidenced by the irregular alignment of immature collagen fibers. Despite the presence of matrix abnormalities, MMP-9-null wounds displayed normal tensile strength. Ultrastructural analysis of wounds revealed the presence of large collagen fibrils, some with irregular shape. Keratinocyte proliferation, inflammation, and angiogenesis were found to be normal in MMP-9-null wounds. In addition, VEGF levels were similar in control and MMP-9-null wound extracts. To investigate the importance of MMP-9 in wound reepithelialization we tested human and murine keratinocytes in a wound migration assay and found that antibody-based blockade of MMP-9 function or MMP-9 deficiency retarded migration. Collectively, our observations reveal defective healing in MMP-9-null mice and suggest that MMP-9 is required for normal progression of wound closure.     

Sulfate Reduction and Methanogenesis in the Sediment of a Saltmarsh on the East Coast of the United Kingdom

The rates of sulfate reduction, methanogenesis, and methane loss were measured in saltmarsh sediment at monthly intervals. In addition, dissolved methane and sulfate concentrations together with pS²⁻ and pH were determined. Methane formation from carbon dioxide, but not from acetate, was detected within the same horizon of sediment where sulfate reduction was most active. Sulfate reduction was about three orders of magnitude greater than annual methanogenesis. The two processes were not separated either spatially or temporally, but occurred within the same layer of sediment at the same time of the year. Their coexistence did not seem to be the result of sulfate-depleted microenvironments within which methanogenesis could occur, but the methanogenic bacteria persisted at very low rates of activity within the same environment as the sulfate reducers.     

The estrogen-receptor complex is bound at unusual chromatin regions

The estrogen receptor of MCF-7 cells labeled with high specific activity estradiol was used to mark the chromatin binding sites for this regulatory molecule. Many of these sites are especially sensitive to nuclease, and produce on digestion a series of uniquely sedimenting products. Several of these have been examined in some detail in this paper. These include a form of receptor that sediments in trace digests at 9S but in more extensive digests at 7S, fast mononucleosomes of about 12.5S, and a species at 15S. Two components of digests, fast mononucleosomes and dinucleosomes were isolated and subjected to further digestion. Much of the hormone on these isolated particles was found to be sensitive to additional hydrolysis, although some was nuclease resistant. It appears that a major fraction of the hormone receptor complexes bound to MCF-7 cell chromatin occurs at nucleosome-free regions which can be detected as transient hydrolysis intermediates.     

Ethnicity as a variable in epidemiological research.

Ethnicity is used increasingly as a key variable to describe health data, and ethnic monitoring in the NHS will further stimulate this trend. We identify four fundamental problems with ethnicity in this type of research: the difficulties of measurement, the heterogeneity of the populations being studied, lack of clarity about the research purpose of the research, and ethnocentricity affecting the interpretation and use of data. Ethnicity needs to be used carefully to be a useful tool for health research. We make nine recommendations for future practice, one of which is that ethnicity and race should be recognised and treated as distinct concepts.     

Functional and structural responses of liver alcohol dehydrogenase to lysine modifications

Acetylation, glycosylation, and methylation, which modify lysine residues of horse liver alcohol dehydrogenase, have been investigated. Acetylation reacted with approximately two-third of the total lysines to induce the greatest structural changes of the enzyme. Glycosylation modified only one lysine residue selectively with indiscernible structural changes. The glycosylation effect was very specific with respect to diastereoisomers for aldopentoses, aldohexoses, and ketohexoses. Methylation produced the largest enhancement in the oxidative activity, which is related to the stability of the modified enzyme to prolonged modification and thermal denaturation. Kinetic studies revealed that a change in the maximal velocity was primarily responsible for the observed activity differences in the modifications.     

Tight divalent metal binding to Escherichia coli F1-adenosinetriphosphatase. Complete substitution of intrinsic magnesium by manganese or cobalt and studies of metal binding sites

Tight divalent metal binding sites in Escherichia coli F1-adenosinetriphosphatase (F1-ATPase) were studied. Native enzyme contained two Mg per F1, confirming previous results. All of the Mg may be replaced by Co or Mn using a dissociation-repolymerization procedure. The substituted enzymes are homogeneous and contain two Mn per F1 or two Co per F1. They are fully active as ATPases, they rebind to F1-depleted membranes, and they catalyze ATP-driven proton pumping. N,N'-Dicyclohexylcarbodiimide-(DCCD) inactivated F1 retains all the intrinsic tightly bound Mg. Evidence is presented that DCCD affects at least two beta subunits in E. coli F1, and therefore, the tightly bound metals appear not to be bound at the DCCD-reactive glutamate residue on the beta subunit. However, the nature of the tightly bound metal (Mg, Mn, or Co) as well as the presence of added (2 mM) MgSO4, MnSO4, or CoSO4 affected the rate of DCCD inactivation, showing that metal binding changes the beta-subunit conformation. Isolated F1 alpha subunit bound Mg, Mn, or Co stoichiometrically and independently of ATP binding. Isolated F1 beta subunit bound only small amounts of Mg, and no Co or Mn. Therefore, it is possible, although not conclusively shown, that the alpha subunit is the site of tight metal binding in the intact F1.     

Dissociation of the chemotactic and mitogenic activities of platelet- derived growth factor by human neutrophil elastase

Because platelet-derived growth factor (PDGF) may be released at sites where neutrophil proteinases may also be released, we examined the effects of neutrophil elastase and cathepsin G upon the chemotactic and mitogenic activities of PDGF. Elastase abolished the chemotactic activity of PDGF for fibroblasts but had no effect on its chemotactic activity for monocytes, or on its mitogenic activity for 3T3 cells or its capacity to bind to 3T3 cells. Cathepsin G had no effect upon the chemotactic or mitogenic activities of PDGF. In contrast, trypsin eliminated the chemotactic activity of PDGF for monocytes and fibroblasts and the mitogenic activity of PDGF. After reduction and alkylation, PDGF retained full chemotactic activity for fibroblasts and monocytes but exhibited no mitogenic activity and only limited binding to 3T3 cells. These results indicate separate domains on PDGF for fibroblast chemotactic and mitogenic activity and for monocyte and fibroblast chemotactic activity and raise the possibility that the biological activities of PDGF may be modified selectively in vivo. The findings further suggest that the majority of PDGF receptors on fibroblasts mediate mitogenic activity and that only a minority of the PDGF receptors on fibroblasts are responsible for chemotactic activity.