Localized transposon Tn5 mutagenesis of the photosynthetic gene cluster of Rhodobacter sphaeroides

Four genes essential for bacteriochlorophyll biosynthesis were known to be encoded within a 45 kb region of the Rhodobacter sphaeroides genome, the boundaries of which are defined by puh and puf genes for reaction-centre and light-harvesting LH1 complexes. The cluster is represented by eight overlapping inserts cloned in the mobilizable vector pSUP202. We have used localized transposon Tn5 mutagenesis to characterize this cluster further; a total of 87 independent insertions were generated which identify nine genes for bacteriochlorophyll biosynthesis, six for carotenoid biosynthesis, and puhA encoding the reaction-centre H subunit. This work provides an essential framework for a detailed study of the structure and expression of genes for photosynthesis in this bacterium.     

The mechanism of phosphoglucomutase from Micrococcus lysodeikticus

The mechanism of the phosphoglucomutase from Micrococcus lysodeikticus was investigated. Induced-transport tests at low substrate concentrations (0.15mm) showed co-transport of the (32)P label but no induced transport of the (14)C label, which is in quantitative agreement with a phosphoenzyme mechanism with a rapid isomerization of the phosphoenzyme. The results excluded an intramolecular transfer of phosphate and could only have been compatible with a sequential mechanism if the K(m) for glucose 1-phosphate had been over 20 times smaller than the measured value. The results of induced-transport tests at intermediate concentrations (1mm) with both labels agreed quantitatively with a phosphoenzyme mechanism, and induced-transport tests with (14)C-labelled substrates at high concentrations (26mm) indicated that the rate constants for isomerization of the phosphoenzyme must be greater than about 3x10(6)s(-1). Consistent with these findings is the fact that (14)C label exchanged between the substrates twice as rapidly as the (32)P label at chemical equilibrium. Further, since the (14)C label exchanged between the substrates about ten times more rapidly than between the substrates and glucose 1,6-diphosphate, glucose 1,6-diphosphate is not an obligatory intermediate in the interconversion of the substrates. It is concluded that, contrary to previous evidence, the mechanism of the enzyme from M. lysodeikticus is essentially that of the rabbit muscle enzyme. To account for the rapid isomerization of the phosphoenzyme in both cases a mechanism is proposed in which there is no formal isomerization of the phosphoenzyme.     

Lifetime consequences of variable 0 year group length in riverine populations of chub Leuciscus cephalus (L.)

The fork length ( L _F) of individual chub Leuciscus cephalus in English riverine populations at the end of their first growth season varied considerably, ranging from <25 to >70 mm. This had a significant influence on the subsequent growth of individuals over their lifetime. Chub of small L _F at age 1 year generally produced smaller annual growth increments throughout life than those of longer L _F at age 1 year, although they had the potential to attain greater ultimate L _F. This variability in L _F at age 1 year resulted, at least in part, from multiple spawning events over a protracted period, that caused variation in the growing seasons for 0 year group chub of the same year class. Since the adult population contained individuals that only attained L _F of <25 mm at the end of their first growing season, it is possible that L _F at age 1 year may not be such a major precursor to strong chub recruitment as previously thought.     

Life cycle and production of Hydrobia acuta Drap. (Gastropoda: Prosobranchia) in a hypersaline coastal lagoon

The life cycle and annual production of Hydrobia acuta was studied in a hypersaline lagoon (s = 39 in summer), forming a part of solar salt works. Quantitative random samples were taken at regular intervals over a period of 15 months using a corer, and snails collected were counted and measured. Weight and biomass was calculated from a length-weight relationship and from measurements of ash content. H. acuta was a strictly annual species in the study lagoon. Recruitment takes place over a brief period in May and June, after which the breeding population dies. Growth of the new generation was slow during summer, probably due to the unfavourably high salinity. A period of rapid growth took place in autumn coinciding with a drop in salinity caused by rainfall. In winter Hydrobia hibernated by burrowing deeply into the sediment. Growth recommenced in spring when the lagoon was reflooded, but by this time the number of survivors was low.The maximum density of snails was 6 000 m^–2 and maximum biomass 500 mg organic dry wt · m^–2. Annual cohort production was estimated as 786 mg organic dry wt · m^–2 · a^–1. These figures are low compared to other studies on hydrobiid snails, and for production in inland waters, but the value for annual P/B = 4.5 is typical for a univoltine species. The relevance of the results to foraging by wading birds (the main consumers), is discussed.     

Mitochondrial carbamoyl phosphate synthetase activity in the absence of N-acetyl-L-glutamate. Mechanism of activation by this cofactor

Rat liver carbamoyl-phosphate synthetase I is shown to have synthetase and ATPase activity in the absence of acetylglutamate. Km values for ATP, Mg2+ and K+ are greatly increased, the Km for HCO-3 is not changed much, and the Km for NH+4 is markedly reduced. Vmax for the synthetase reaction is less than 20% of that of the acetylglutamate-activated enzyme whereas Vmax for the ATPase activity is greater than 40% of that with acetylglutamate. Pulse-chase experiments with H14CO-3 show formation of less "active CO2" (the central intermediate) than with acetylglutamate; ATPase activity is reduced in proportion, but the synthetase activity is much smaller. Binding of one ATP molecule with high affinity (Kd = 20-30 microM) is shown in the absence of acetylglutamate. This appears to be the molecule of ATPB (ATPB provides the phosphoryl group of carbamoyl phosphate). In contrast, the affinity for ATPA (ATPA yields Pi) is much reduced. Initial velocity measurements without acetylglutamate show a time lag before reaching a constant velocity. At 50 microM acetylglutamate the lag is much longer, but at 10 mM acetylglutamate it is shorter. Activation by acetylglutamate requires ATP at concentrations sufficient to occupy the ATPA and the ATPB binding sites. Preincubation with 10 mM acetylglutamate alone shortens the activation time. From these findings we propose an allosteric model for activation of carbamoyl-phosphate synthetase in which there are two active states, R and R . AcGlu. Binding of ATPA is associated with the conversion of T to R. R . AcGlu differs from R in that transfer to carbamate of the gamma-phosphoryl group of ATPB appears to be facilitated.     

A singular dispersion relation arising in a caricature of a model for morphogenesis

In 1983 Oster et al. proposed a model for morphogenesis consisting of a system of partial differential equations in which the dispersion relation for the problem linearised about the zero solution has a singularity. That is, the initial growth rate of a small perturbation of wave number k from the zero solution tends to positive or negative infinity as k tends to some critical value k _c from above or below respectively. We consider here as a caricature of the model a single partial differential equation with a similar dispersion relation in a bounded one-dimensional domain. The wave number, or equivalently the domain size, may be thought of as a bifurcation parameter. For the Neumann problem a phenomenon arises in which, as the domain size l increases past a critical value l _l ,the linear stability of the n-th mode jumps from one solution to a remote solution. That is, for l _n the trivial solution is unstable and a certain non-trivial solution is stable to perturbations of mode n, whereas for l>l _n the opposite is true. For the Dirichlet or the Robin problem a linear stability change in the trivial solution occurs, but no corresponding change in any other solution has been found. The corresponding initial boundary value problems are then considered. An asymptotic analysis is performed in the weakly nonlinear limit in the particular case in which only one mode is unstable and gives an asymptotic solution for two classes of nonlinearity, one symmetric and the other asymmetric about u=0. A development of the method of harmonic balance is then used to obtain approximate solutions in the strongly nonlinear case and when more than one mode may be unstable.     

Chlordecone does not interfere with hepatic repair after carbon tetrachloride or partial hepatectomy

The effect of chlordecone (CD) on hepatic repair, measured either as recovery of microsomal enzymatic functions or as the induction of cytosolic thymidine kinase (TK) activity, was evaluated in rats given carbon tetrachloride (CC1₄). Carbon tetrachloride was administered to CD-potentiated and control animals using doses of this hepatotoxin which produce similar degrees of damage at 24 hours in both groups of animals (6 and 100 μ1 CC1₄ per 100 g body weight, respectively). Chlordecone had no significant effect on the time course of recovery of microsomal cytochrome P-450 content or glucose-6-phosphatase activity following CC1₄ administration. Hepatic TK activity was measured 48 hours after CC1₄ administration as a biochemical index of the hepatic regenerative response. Thymidine kinase activity was increased eightfold in CD-treated rats receiving 6 μ1 CC1₄ per 100 g body weight, whereas in controls a similar induction of TK activity was produced by 100 μ1 CC1₄ per 100 g body weight. Therefore, the TK response in CD-treated rats receiving CC1₄ is appropriate for the amount of damage produced, suggesting that CD does not inhibit the hepatic regenerative response to CC1₄-induced injury. The effect of CD on hepatic repair was also examined in rats receiving a two-thirds partial hepatectomy. Pretreatment of animals with CD had no significant effect on the increase in TK activity produced 24 hours after partial hepatectomy. These results offer no support for the idea that CD impairs hepatic repair after either partial hepatectomy or CC1₄ administration.     

Potentiation of carbon tetrachloride hepatotoxicity by chlordecone: Dose-response relationships and increased covalent binding in vivo

Chlordecone greatly potentiates carbon tetrachloride (CC1₄) hepatotoxicity. In order to quantitate the degree of this potentiation, the effects of a range of doses of CC1₄ on two microsomal enzymatic functions and liver enzyme release were examined in chlordecone-treated and control rats. Male Sprague-Dawley rats were pretreated with 15 mg chlordecone per kilogram body weight (BW) intragastrically or with vehicle. After 48 hours, 0 to 250 μ1 CC1₄ per 100 g body weight were given intraperitoneally (IP), and the rats were killed 24 hours later. Chlordecone treatment produced approximately a 17-fold potentiation of the CC1₄ dependent loss of cytochrome P-450 and glucose-6-phosphatase activity, so that a dose of 6 μ1 CC1₄ per 100 g body weight in the chlordecone-treated animals resulted in a similar amount of damage as observed with 100 μ1 CC1₄ per 100 g body weight in controls. A similar potentiation by chlordecone was seen with CC1₄- induced increases in serum glutamic-oxaloacetic transaminase (SGOT) levels. Chlordecone treatment also increased hepatic cytochrome P-450 levels by 67% and resulted in an increase in the covalent binding of [14-C]-CC1₄-derived metabolites to microsomal protein and lipid in vivo.