Tight ATP and ADP binding in the noncatalytic sites of Escherichia coli F1-ATPase is not affected by mutation of bulky residues in the 'glycine-rich loop'

It is shown that ATP dissociates very slowly (koff less than 6.4 x 10(5) s-1, t1/2 greater than 3 h) from the three noncatalytic sites of E. coli F1-ATPase and that ADP dissociates from these three sites in a homogeneous fashion with koff = 1.5 x 10(-4) s-1 (t1/2 = 1.35 h). Mutagenesis of alpha-subunit residues R171 and Q172 in the 'glycine-rich loop' (Homology A) consensus region of the noncatalytic sites was carried out to test the hypothesis that unusually bulky residues at these positions are responsible wholly or partly for the observed tight binding of adenine nucleotides. The mutations alpha Q172G or alpha R171S,Q172G had no effects on ATP or ADP binding to or rates of dissociation from F1 noncatalytic sites. KdATP and KdADP of isolated alpha-subunit were weakened by approximately 1 order of magnitude in both mutants. The results suggest that neither residue alpha R171 nor alpha Q172 interacts directly with bound nucleotide, and show that the presence of bulky residues per se in the glycine-rich loop region of F1-alpha-subunit is not responsible for tight binding in the noncatalytic sites.     

Tropical forests are thermally buffered despite intensive selective logging

Tropical rainforests are subject to extensive degradation by commercial selective logging. Despite pervasive changes to forest structure, selectively logged forests represent vital refugia for global biodiversity. The ability of these forests to buffer temperature‐sensitive species from climate warming will be an important determinant of their future conservation value, although this topic remains largely unexplored. Thermal buffering potential is broadly determined by: (i) the difference between the “macroclimate” (climate at a local scale, m to ha) and the “microclimate” (climate at a fine‐scale, mm to m, that is distinct from the macroclimate); (ii) thermal stability of microclimates (e.g. variation in daily temperatures); and (iii) the availability of microclimates to organisms. We compared these metrics in undisturbed primary forest and intensively logged forest on Borneo, using thermal images to capture cool microclimates on the surface of the forest floor, and information from dataloggers placed inside deadwood, tree holes and leaf litter. Although major differences in forest structure remained 9–12 years after repeated selective logging, we found that logging activity had very little effect on thermal buffering, in terms of macroclimate and microclimate temperatures, and the overall availability of microclimates. For 1°C warming in the macroclimate, temperature inside deadwood, tree holes and leaf litter warmed slightly more in primary forest than in logged forest, but the effect amounted to <0.1°C difference between forest types. We therefore conclude that selectively logged forests are similar to primary forests in their potential for thermal buffering, and subsequent ability to retain temperature‐sensitive species under climate change. Selectively logged forests can play a crucial role in the long‐term maintenance of global biodiversity.     

An evaluation of the utility of SSR loci as molecular markers in maize (Zea mays L.): comparisons with data from RFLPS and pedigree

 The utility of 131 simple sequence repeat (SSR) loci to characterize and identify maize inbred lines, validate pedigree, and show associations among inbred lines was evaluated using a set of 58 inbred lines and four hybrids. Thirteen sets of inbred parent-progeny triplet pedigrees together with four hybrids and their parental lines were used to quantify incidences of scoring that departed from expectations based upon simple Mendelian inheritance. Results were compared to those obtained using 80 restriction fragment length polymorphism (RFLP) probes. Over all inbred triplets, 2.2% of SSRs and 3.6% of RFLP loci resulted in profiles that were scored as having segregated in a non-Mendelian fashion. Polymorphic index content (PIC, a measure of discrimination ability) values ranged from 0.06 to 0.91 for SSRs and from 0.10 to 0.84 for RFLPs. Mean values for PIC for SSRs and RFLPs were similar, approximately 0.62. However, PIC values for nine SSRs exceeded the maximum PIC for RFLPs. Di-repeats gave the highest mean PIC scores for SSRs but this class of repeats can result in “stutter” bands that complicate accurate genotyping. Associations among inbreds were similar for SSR and RFLP data, closely approximating expectations from known pedigrees. SSR technology presents the potential advantages of reliability, reproducibility, discrimination, standardization and cost effectiveness over RFLPs. SSR profiles can be readily interpreted in terms of alleles at mapped loci across a broad range of maize germ plasm. Consequently, SSRs represent the optimum approach for the identification and pedigree validation of maize genotypes compared to other currently available methods. Received: 15 January 1997 / Accepted: 28 February 1997     

16-Aryloxyprostaglandins: A new class of potent luteolytic agent

Recent work has established that chloroform extracts of ram semen and fractions of these extracts accelerate rejection of the nematode, Nippostrongylus brasiliensis, from the intestine of rats when injected intra-duodenally on day 6 of a primary infection (6). It was also shown that the administration of aspirin and d-propoxyphene hydrochloride (d-PP), potent inhibitors of prostaglandin action (7, 8), prevented the expulsion of worms which normally occurs between days 10 and 16 of a primary infection with N. brasiliensis. In the present study, we have established that there is a direct correlation between smooth muscle contracting activity and the capacity of individual semen fractions to accelerate worm expulsion. Methylation destroyed both smooth muscle contracting activity and the capacity of semen fractions to cause worm expulsion. Contraction of smooth muscle induced by the most active semen fraction (S.A.F. 1) was not inhibited by the amine antagonists mepyramine maleate and bromylsergic acid diethylamide. In addition, contractions induced in rabbit duodenum segments by 5-hydroxytryptamine were not inhibited by aspirin. These findings indicate that the semen fractions do not contain physiologically significant levels of the amines, histamine and 5-hydroxytryptamine, and this suggests that the capacity of semen fractions to cause worm expulsion is due to prostaglandins. This conclusion is supported by the observation that the most active fraction S.A.F. 1 contained bands with R_F values which corresponded with the R_F values of synthetic prostaglandins in thin layer chromatography. Furthermore, the intra-duodenal injection of synthetic prostaglandins also caused worm expulsion.     

Light-scattering and potentiometric-titration studies of poly-L-tyrosine in aqueous solution

Aqueous solutions of poly-L -tyrosine were studied by potentiometric titrations, light-scattering measurements, and infrared spectroscopy in order to characterize the conformational changes the macromolecules exhibit when the hydrogen-ion concentration of the solutions is varied. It was found that when the pH of the system at 25°C is lowered from a value of about 13 the poly-L -tyrosine molecules undergo a random-coil to β-structure conformation change at pH 11.5. It appears, that under the experimental conditions used, the formation of the β structure is an intramolecular process which involves the folding of the polymer backbone into several hairpins of antiparallel β structure. On further lowering of the pH of the solutions aggregation (pH 11.35) and subsequently precipitation (pH 11.2) takes place. Since the apparent pK of the polymer does not show a drastic change during the random-coil to β-structure conformation change, it was concluded, that the therinodynamic driving force for this conformation change is mainly due to the electrostatic effect of the partially charged side chains of the poly-L -tyrosine molecules.     

Biochemical effects of the hypoglycaemic compound pent-4-enoic acid and related non-hypoglycaemic fatty acids. Oxidative phosphorylation and mitochondrial oxidation of pyruvate, 3-hydroxybutyrate and tricarboxylic acid-cycle intermediates

1. The effects of the hypoglycaemic compound pent-4-enoic acid, and of four structurally related non-hypoglycaemic compounds (pent-2-enoic acid, pentanoic acid, cyclopropanecarboxylic acid and cyclobutanecarboxylic acid), on several reactions in rat liver mitochondria were determined. 2. The use of manometric techniques for measurements of oxidations and of phosphorylation is critically discussed. 3. Pent-4-enoic acid and pentanoic acid uncoupled oxidative phosphorylation at low concentrations, but usually by not more than about 50%. 4. All the compounds, except cyclobutanecarboxylic acid, strongly inhibited the oxidation of pyruvate and 2-oxoglutarate, but the oxidations of succinate, citrate and 3-hydroxybutyrate were not strongly inhibited. 5. All the compounds, except cyclobutanecarboxylic acid, inhibited decarboxylation of [1-(14)C]pyruvate with ferricyanide as electron acceptor. 6. All the compounds, except pent-2-enoic acid, caused mitochondrial swelling after a time-lag.     

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.