Thermal denaturation and loss of viability in Escherichia coli and Bacillus stearothermophilus

When Escherichia coli was heated at 10°C/min in a differential scanning calorimeter, the onset of irreversible thermal denaturation occurred at 51°C, about 5°C above the maximum growth temperature. The temperature at which death rate was maximal (63°C) coincided with the thermogram peak caused by denaturation of the 30S ribosomal subunit. The maximum death rate in vegetative cells of Bacillus stearothermophilus occurred at the higher temperature of 71°C which also coincided with the leading edge of the main thermogram peak.     

Identification and sequence analysis of cDNAs encoding a 110-kilodalton actin filament-associated pp60src substrate.

Transformation of chicken embryo cells by oncogenic forms of pp60src (e.g., pp60v-src or pp60527F) is linked with a concomitant increase in the steady-state levels of tyrosine-phosphorylated cellular proteins. Activated forms of the Src protein-tyrosine kinase stably associate with tyrosine-phosphorylated proteins, including a protein of 110 kDa, pp110. Previous reports have established that stable complex formation between pp110 and pp60src requires the structural integrity of the Src SH2 and SH3 domains, whereas tyrosine phosphorylation of pp110 requires only the structural integrity of the SH3 domain. In normal chicken embryo cells, pp110 colocalizes with actin stress filaments, and in Src-transformed cells, pp110 is found associated with podosomes (rosettes). Here, we report the identification and characterization of cDNAs encoding pp110. The predicted open reading frame encodes a polypeptide of 635 amino acids which exhibits little sequence similarity with other protein sequences present in the available sequence data bases. Thus, pp110 is a distinctive cytoskeleton-associated protein. On the basis of its association with actin stress filaments, we propose the term AFAP-110, for actin filament-associated protein of 110 kDa. In vitro analysis of AFAP-110 binding to bacterium-encoded glutathione S-transferase (GST) fusion proteins revealed that AFAP-110 present in normal cell extracts binds efficiently to Src SH3/SH2-containing fusion proteins, less efficiently to Src SH3-containing proteins, and poorly to SH2-containing fusion proteins. In contrast, AFAP-110 in Src-transformed cell extracts bound to GST-SH3/SH2 and GST-SH2 fusion proteins. Analysis of AFAP-110 cDNA sequences revealed the presence of sequence motifs predicted to bind to SH2 and SH3 domains, respectively. We suggest that AFAP-110 may represent a cellular protein capable of interacting with SH3-containing proteins and, upon tyrosine phosphorylation, binds tightly to SH2-containing proteins, such as pp60src or pp59fyn. The potential roles of AFAP-110 as an SH3/SH2 cytoskeletal binding protein are discussed.     

The unexpected long period of elevated CH4 emissions from an inundated fen meadow ended only with the occurrence of cattail (Typha latifolia)

Drainage and agricultural use transform natural peatlands from a net carbon (C) sink to a net C source. Rewetting of peatlands, despite of high methane (CH₄) emissions, holds the potential to mitigate climate change by greatly reducing CO₂ emissions. However, the time span for this transition is unknown because most studies are limited to a few years. Especially, nonpermanent open water areas often created after rewetting, are highly productive. Here, we present 14 consecutive years of CH₄ flux measurements following rewetting of a formerly long-term drained peatland in the Peene valley. Measurements were made at two rewetted sites (non-inundated vs. inundated) using manual chambers. During the study period, significant differences in measured CH₄ emissions occurred. In general, these differences overlapped with stages of ecosystem transition from a cultivated grassland to a polytrophic lake dominated by emergent helophytes, but could also be additionally explained by other variables. This transition started with a rapid vegetation shift from dying cultivated grasses to open water floating and submerged hydrophytes and significantly increased CH₄ emissions. Since 2008, helophytes have gradually spread from the shoreline into the open water area, especially in drier years. This process was periodically delayed by exceptional inundation and eventually resulted in the inundated site being covered by emergent helophytes. While the period between 2009 and 2015 showed exceptionally high CH₄ emissions, these decreased significantly after cattail and other emergent helophytes became dominant at the inundated site. Therefore, CH₄ emissions declined only after 10 years of transition following rewetting, potentially reaching a new steady state. Overall, this study highlights the importance of an integrative approach to understand the shallow lakes CH₄ biogeochemistry, encompassing the entire area with its mosaic of different vegetation forms. This should be ideally done through a study design including proper measurement site allocation as well as long-term measurements.     

Organization of the genes necessary for hydrogenase expression in Rhodobacter capsulatus. Sequence analysis and identification of two hyp regulatory mutants

A 25kbp DNA fragment from the chromosome of Rhodobacter capsulatus B10 carrying hydrogenase (hup) determinants was completely sequenced. Coding regions corresponding to 20 open reading frames were identified. The R. capsulatus hydrogenase-specific gene (hup and hyp) products bear significant structural identity to hydrogenase gene products from Escherichia coli (13), from Rhizobium liguminosarum (16), from Azotobacter vinelandii (10) and from Alcaligenes eutrophus (11). The sequential arrangement of the R. capsulatus genes is: hupR2-hupU-hypF -hupS-hupL-hupM-hupD -hupF -hupG -hupH -huoJ -hupK -hypA-hypB-hupR1-hypC -hypD -hypE -ORF19 -ORF20 , all contiguous and transcribed from the same DNA strand. The last two potential genes do not encode products that are related to identified hydrogenase-specific gene products in other species. The sequence of the 12 R. capsulatus genes underlined above is presented. The mutation site in two of the Hup^? mutants used in this study, RS13 and RCC12, was identified in the hypF gene (deletion of one G) and in the hypD qene (deletion of 54 bp), respectively. The hypF gene product shares 45% identity with the product of hydA from E. coli and the product of hypF from R. leguminosarum. Those products present at their N-terminus a Cys arrangement typical of zinc-finger proteins. The G deletion in the C-terminal region of hypF in the RS13 mutant     

Differentiation of species in Gonioloboceras

The type specimen ofGonioloboceras goniolobum (Meek), rediscovered by Spath in the British Museum, is the foundation for a more accurate comparative study of this and other species ofGonioloboceras.Gonioloboceras described asG. goniolobum byElias in 1938 is differentiated asGonioloboceras schmidti, new species. Suture sets (new term) for several growth stages inG. goniolobum (Meek),G. welleriSmith,G. schmidtiElias, G.eliasiMiller &Owen, andG. asiaticumLibrovitch are assembled and used for differentiation of the species.The Kazakhstan goniatite faunule containingG. asiaticum is considered of very late Pennsylvanian age.     

Aerobic degradation of polychlorinated biphenyls by Alcaligenes sp. JB1: metabolites and enzymes

In contrast to the degradation of penta-and hexachlorobiphenyls in chemostat cultures, the metabolism of PCBs by Alcaligenes sp. JB1 was shown to be restricted to PCBs with up to four chlorine substituents in resting-cell assays. Among these, the PCB congeners containing ortho chlorine substituents on both phenyl rings were found to be least degraded. Monochloro-benzoates and dichlorobenzoates were detected as metabolites. Resting cell assays with chlorobenzoates showed that JB1 could metabolize all three monochlorobenzoates and dichlorobenzoates containing only meta and para chlorine substituents, but not dichlorobenzoates possessing an ortho chlorine substituent. In enzyme activity assays, meta cleaving 2,3-dihydroxybiphenyl 1,2-dioxygenase and catechol 2,3-dioxygenase activities were constitutive, whereas benzoate dioxygenase and ortho cleaving catechol 1,2-dioxygenase activities were induced by their substrates. No activity was found for pyrocatechase II, the enzyme that is specific for chlorocatechols. The data suggest that complete mineralization of PCBs with three or more chlorine substituents by Alcaligenes sp. JB1 is unlikely.Abbreviations PCB polychlorinated biphenyls - CBA chlorobenzoate - D di - Tr tri - Te tetra - Pe penta- - H hexa     

Ecological compensation of the impacts of a road. Preliminary method for the A50 road link (Eindhoven-Oss, The Netherlands)

After years of efforts to avoid or reduce the impact of infrastructure projects on nature, the principle of ecological compensation has been incorporated in Dutch governmental policy. Ecological compensation aims to recover those ecological functions and natural values that still remain affected after maximum effort has been made to reduce the impact of the intervention (mitigation). The accepted aim of current policy is thus no-net-loss of area and quality by means of mitigative and compensatory measures. As part of the planning process for construction of a stretch of road in The Netherlands, viz., the A50 road link in the province of North Brabant, a Nature Compensation Plan (NCP) was required to be drawn up. This work has recently been completed by the Regional Directorate of Public Works and Water Management, the initiator of the intervention. The NCP, initially presented as a Draft Plan, was drawn up by the Regional Directorate using a preliminary method designed by the Centre of Environmental Science of Leiden University for deriving compensatory measures. After an opportunity for public comment, the Draft Plan was revised to form a Final NCP. This article describes, firstly, the preliminary method for deriving ecological compensatory measures. The method starts by quantifying the effects of habitat loss, habitat disturbance (by changes in noise emissions, in the water table and in outdoor recreational patterns), barrier action and fauna casualties. Following mitigation of impacts on nature, compensation for non-mitigable effects focuses successively on area size, derived from the impacts on breeding birds, and on area quality, derived from the habitat requirements of the vegetation and fauna groups affected by the road. Guidelines for identifying appropriate locations for compensation are also formulated. Secondly, the compensation method is applied to calculate the mitigative and compensatory measures for the A50 trajectory between Eindhoven and Oss. Thirdly, two comparisons are made: the Draft NCP is compared with the results of the preliminary method, and the Draft and the Final NCP are compared with one another in order to identify the role of the interest groups that played a major role in commenting on the Draft Plan. Finally, realization of the compensatory measures and development of the preliminary method itself are discussed. On the basis of the experience with the A50 case study, a more robust compensation method for road projects is to be developed.     

A third family of allelic hsd genes in Salmonella enterica: sequence comparisons with related proteins identify conserved regions implicated in restriction of DNA

Salmonella enterica serovar blegdam has a restriction and modification system encoded by genes linked to serB . We have cloned these genes, putative alleles of the hsd locus of Escherichia coli  K-12, and confirmed by the sequence similarities of flanking DNA that the hsd genes of S. enterica serovar blegdam have the same chromosomal location as those of E. coli K-12 and Salmonella enterica serovar typhimurium LT2. There is, however, no obvious similarity in their nucleotide sequences, and while the gene order in S. enterica serovar blegdam is serB hsdM , S and R , that in E. coli K-12 and S. enterica serovar typhimurium LT2 is serB hsdR , M and S . The hsd genes of S. enterica serovar blegdam identify a third family of serB -linked hsd genes (type ID). The polypeptide sequence predicted from the three hsd genes show some similarities (18–50% identity) with the polypeptides of known and putative type I restriction and modification systems; the highest levels of identity are with sequences of Haemophilus influenzae Rd. The HsdM polypeptide has the motifs characteristic of adenine methyltransferases. Comparisons of the HsdR sequence with those for three other families of type I systems and three putative HsdR polypeptides identify two highly conserved regions in addition to the seven proposed DEAD-box motifs.