Microbial growth on carbon monoxide

The utilization of carbon monoxide as energy and/or carbon source by different physiological groups of bacteria is described and compared. Utilitarian CO oxidation which is coupled to the generation of energy for growth is achieved by aerobic and anaerobic eu- and archaebacteria. They belong to the physiological groups of aerobic carboxidotrophic, facultatively anaerobic phototrophic, and anaerobic acetogenic, methanogenic or sulfate-reducing bacteria. The key enzyme in CO oxidation is CO dehydrogenase which is a molybdo iron-sulfur flavoprotein in aerobic CO-oxidizing bacteria and a nickel-containing iron-sulfur protein in anaerobic ones. In carboxidotrophic and phototrophic bacteria, the CO-born CO₂ is fixed by ribulose bisphosphate carboxylase in the reductive pentose phosphate cycle. In acetogenic, methanogenic, and probably in sulfate-reducing bacteria, CODH/acetyl-CoA synthase directly incorporates CO into acetyl-CoA.In plasmid-harbouring carboxidotrophic bacteria, CO dehydrogenase as well as enzymes involved in CO₂ fixation or hydrogen utilization are plasmid-encoded. Structural genes encoding CO dehydrogenase were cloned from carboxidotrophic, acetogenic and methanogenic bacteria. Although they are clustered in each case, they are genetically distinct.Soil is a most important biological sink for CO in nature. While the physiological microbial groups capable of CO oxidation are well known, the type and nature of the microorganisms actually representing this sink are still enigmatic. We also tried to summarize the little information available on the nutritional and physicochemical requirements determining the sink strength. Because CO is highly toxic to respiring organisms even in low concentrations, the function of microbial activities in the global CO cycle is critical.     

Substance P and enkephalin in transected axons of medulla and spinal cord

The accumulation of transported materials in cut axons is demonstrated by the light and electron microscopic immunocytochemical localization of substance P and enkephalin in the caudal medulla and cervical spinal cord of adult rat. Two days following unilateral knife-cuts in the caudal medulla or spinal (C2-C3) levels, substance P and enkephalin-like immunoreactivity (SPLI and ELI) are detected in lesioned axons located rostral and caudal to the transection. Rostrally, SPLI and ELI are detected in the lateral reticular region and ventrolateral fasciculus corresponding to the location of previously identified bulbospinal pathways. Caudally, previously unidentified, propriospinal pathways showing SPLI are detected in the dorsal columns and in the dorsolateral fasciculus. In contrast, ELI is found caudal to the transection only in the reticular region of the medulla. For both peptides, immunoreactivity is present throughout axons containing numerous large, dense core, and small clear vesicles. These results support the concept of both particulate and soluble modes of transport for substance P and enkephalin within axons of the central nervous system.     

Causal connection between detoxification enzyme activity and consumption of a toxic plant compound

Insect herbivores can increase their detoxification activities against a particular plant poison in response to prolonged ingestion of the same compound. For example, larval tobacco hornworms (Manduca sexta) experience a dramatic increase in cytochrome P450 activity against nicotine after ingesting nicotine. While it is generally assumed that this induction process permits increased consumption of toxic plant tissues, we are not aware of any direct experimental support for this assumption. Using a two-tiered approach, we examined the functional significance of P450 induction to M. sexta larvae ingesting a toxic but non-deterrent concentration of nicotine. First, we related the time-course of P450 induction in midgut microsomes to changes in nicotine consumption. When offered a nicotine diet, larvae failed to show a significant increase in consumption before 36 h, which was coincident with the time-course of the induction of midgut P450 activities against aldrin and nicotine. Second, we determined whether inhibiting the induced P450 activities affected nicotine consumption. We found that the increase in nicotine consumption following the induction of nicotine metabolism could be strongly inhibited by treatment with piperonyl butoxide, which by itself did not inhibit consumption. These results provide direct evidence for a causal connection between P450-mediated detoxification activity and consumption of a toxic plant compound.Abbreviation PB piperonyl butoxide     

Catabolism of caffeine and related purine alkaloids in leaves ofCoffea arabica L.

In a study of purine alkaloid catabolism pathways in coffee,¹⁴C-labelled theobromine, caffeine, theophylline and xanthine were incubated with leaves ofCoffea arabica. Incorporation of label into¹⁴CO₂ was determined and methanol-soluble metabolites were analysed by high-performance liquid chromatography-radiocounting. The data obtained demonstrate catabolism of caffeine theophylline 3-methylxanthine xanthine. Xanthine is degraded further by the conventional purine catabolism pathway to CO₂ and NH₃ via uric acid, allantoin and allantoic acid. The conversion of caffeine to theophylline is the rate-limiting step in purine alkaloid catabolism and provides a ready explanation for the high concentration of endogenous caffeine found inC. arabica leaves. Although theobromine is converted primarily to caffeine, a small portion of the theobromine pool appears to be degraded to xanthine by a caffeine-independent pathway. In addition to being broken down to CO₂, via the purine catabolism pathway, xanthine is metabolised to 7-methylxanthine. Metabolism of [2-¹⁴C]xanthine byC. arabica leaves in the presence of 5 mM allopurinol results in very large increases in incorporation of radioactivity into 7-methylxanthine as degradation of the substrate via the purine catabolism pathway is blocked. The identity of 7-methylxanthine in these studies was confirmed by gas chromatography-mass spectrometry analysis.Abbreviations HPLC-RC high-performance liquid chromatography-radiocounting This work was supported by the British Council which provided H.A. with Japan-UK travel grants. F.M.G. was supported by a Biotechnology and Biological Sciences Research Council grant to A.C.     

Hydrochlorothiazide enhances the apical Cl− backflux in rabbit gallbladder epithelium: Radiochemical analysis

Hydrochlorothiazide (HCTZ) was shown to inhibit the transepithelial NaCl transport and the apical Na⁺-Cl^? symport and to depolarize the apical membrane potential in the rabbit gallbladder epithelium. The depolarization was likely related to the opening of a Cl^? conductance. To better understand whether an apical Cl^? leak is involved in the mechanism of action of HCTZ, the transapical Cl^? backflux was measured radiochemically by the washout technique. The gallbladder wall, pretreated with pronase on the serosal side to homogenize the subepithelium, was loaded with ³⁶Cl^? on the luminal side; mucosal and serosal ³⁶Cl^? effluxes (J _m , J _s ) were then measured every 2 min. The pretreatment with pronase did not alter the membrane potentials and the selectivity of the epithelium. Under control conditions and the tissue in steady-state, J _m and J _s time courses were each described by two exponential decays (A,B); the rate constants, k ^A and k ^B , were 0.71 ±0.03 and 0.16±0.01 min^?1, respectively, and correspondingly the half-times (t ₁ ^2A , t ₁ ^2B ) were 1.01±0.05 and 5.00±0.44 min (n=10); these parameters were not significantly different for J _m and J _s time courses. J _s was always greater than J _m (J _s /J _m =2.02±0.22 and 1.43 ±0.17 for A and B decays). Under SCN^? treatment in steady-state conditions, both J _m and J _s time courses were described by only one exponential decay, the component B being abolished. Moreover t ₁ ^2A was similar to that predictable for the subepithelium. It follows that it is the component B which exits the epithelial compartment. Based on the intracellular specific activity and ³⁶Cl^? J _m ^B at 0 min time of the washout experiment, the cell-lumen Cl^? backflux in steady-state was calculated to be equal to about 2 μmol cm^?2hr^?1, in agreement with the value indirectly computable by other techniques. The experimental model was well responsive to different external challenges (increases in media osmolalities; luminal treatment with nystatin). HCTZ (2.5 · 10^?4 m) largely increased ³⁶Cl^? J _m ^B . The increase was abolished by luminal treatment with 10^?4 m SITS, which not only brought back the efflux time courses to the ones observed under control conditions but even increased J _s /J _m of the cellular component, an indication of a reduced J _m ^B . It is concluded that HCTZ opens an apical, SITS-sensitive Cl^? leak, which contributes to dissipate the intracellular Cl^? accumulation and to inhibit the NaCl transepithelial transport. Moreover, the drug is likely to reduce the basal electroneutral Cl^? backflux supported by Na⁺-Cl^? cotransport, in agreement with the inhibition of the cotransport itself.     

Monte carlo simulations of protein folding. II. Application to protein A,ROP, and crambin

The hierarchy of lattice Monte Carlo models described in the accompanying paper (Kolinski, A., Skolnick, J. Monte Carlo simulations of protein folding. I. Lattice model and interaction scheme. Proteins 18:338–352, 1994) is applied to the simulation of protein folding and the prediction of 3-dimensional structure. Using sequence information alone, three proteins have been successfully folded: the B domain of staphylococcal protein A, a 120 residue, monomeric version of ROP dimer, and crambin. Starting from a random expanded conformation, the model proteins fold along relatively well-defined folding pathways. These involve a collection of early intermediates, which are followed by the final (and rate-determining) transition from compact intermediates closely resembling the molten globule state to the native-like state. The predicted structures are rather unique, with native-like packing of the side chains. The accuracy of the predicted native conformations is better than those obtained in previous folding simulations. The best (but by no means atypical) folds of protein A have a coordinate rms of 2.25 Å from the native Cα trace, and the best coordinate rms from crambin is 3.18 Å. For ROP monomer, the lowest coordinate rms from equivalent Cαs of ROP dimer is 3.65 Å. Thus, for two simple helical proteins and a small α/β protein, the ability to predict protein structure from sequence has been demonstrated. © 1994 John Wiley & Sons, Inc.     

The evolution of pathways for aromatic hydrocarbon oxidation inPseudomonas

The organisation and nucleotide sequences coding for the catabolism of benzene, toluene (and xylenes), naphthalene and biphenylvia catechol and the extradiol (meta) cleavage pathway inPseudomonas are reviewed and the various factors which may have played a part in their evolution are considered. The data suggests that the complete pathways have evolved in a modular way probably from at least three elements. The commonmeta pathway operons, downstream from the ferredoxin-like protein adjacent to the gene for catechol 2,3-dioxygenase, are highly homologous and clearly share a common ancestry. This common module may have become fused to a gene or genes the product(s) of which could convert a stable chemical (benzoate, salicylate, toluene, benzene, phenol) to catechol, thus forming the lower pathway operons found in modern strains. The upper pathway operons might then have been acquired as a third module at a later stage thus increasing the catabolic versatility of the host strains.     

Cell cycle mutation in Paramecium tetraurelia discriminates between sexual and vegetative functions

The temperature-sensitive mutation cc1 blocks a number of cell cycle processes in Paramecium including macronuclear DNA synthesis, oral morphogenesis, and the later stages of micronuclear mitosis. Oral morphogenesis and micronuclear mitosis also occur in the sexual pathway. This study shows that cc1 cells can proceed through conjugation or autogamy under restrictive conditions; neither stomatogenesis nor micronuclear mitosis is blocked. Fertilization and macronuclear determination occur normally, but DNA synthesis in macronuclear anlagen is blocked. Therefore, this mutation discriminates between oral replacement during meiosis and vegetative prefission stomatogenesis, and between mitotic spindle elongation during the pregamic and postzygotic divisions and spindle elongation during the vegetative cell cycle. These results point to a fundamental regulatory difference between morphogenesis in the vegetative and sexual pathways. © 1994 Wiley-Liss, Inc.     

Membrane-delimited cell signaling complexes: Direct ion channel regulation by G Proteins

Summary Ion channels are signaling molecules and by them-selves perform no work. In this regard they are un like the usual membrane enzyme effectors for G proteins. The pathways of G protein receptor, G protein and ion channels are, therefore, purely infor mational in function. Because a single G protein may have several ion channels as effectors, the effects should be coordinated and this seems to be the case. Inhibition of Ca²⁺ current and stimulation of K⁺ currents would have a greater impact than either alone. Additional flexibility is provided by spontane ous noise in the complexes of G protein receptor, G protein, and ion channel. By having a non-zero setpoint, the range of control is extended and the responses become bi-directional.     

Evidence for a GABAergic Projection from the Dorsal Nucleus of the Lateral Lemniscus to the Inferior Colliculus

Abstract: This study attempts to determine whether the pathways from the guinea pig dorsal nucleus of the lateral lemniscus (DNLL) to the inferior colliculus (IC) use γ-aminobutyric acid (GABA) as a transmitter. Injections of kainic acid (KA) were used to destroy neurons in the left DNLL. Two to 4 days after the injection, Nissl-stained sections through the lesion site showed destruction of the DNLL neurons. The lesions varied in size; 12–100% of the DNLL neurons were destroyed on the injected side without damage to the ipsilateral IC. Two to 4 days after the injection, the electrically evoked, Ca²⁺-dependent release and high-affinity uptake of [³H]GABA were measured in dissected pieces of the left and right IC. These activities were compared with those in the IC taken from unlesioned controls and from sham controls, which received injections of saline instead of KA. Each IC was divided into a dorsal piece, which contained the dorsal cortex and dorsomedial nucleus, and a ventral piece, which contained the central and lateral nuclei. Lesions of the left DNLL depressed the release and uptake of [³H]GABA in the ventral pieces of the IC, but there was a greater depression in the ventral IC contralateral to the lesioned DNLL. There were good correlations between the percentage of neuronal loss in the left DNLL and deficits in [³H]GABA release and uptake activities in the ipsi- and contralateral ventral IC. By contrast, there was no depression of [³H]GABA release and uptake in the dorsal pieces of the IC. The localization of the deficits in release and uptake appears to match the distribution of the synaptic endings of the DNLL pathways in the IC. This correspondence associates GABA release and uptake activities with the DNLL projections to the IC and, therefore, suggests that GABA may be a transmitter of these pathways. The release and uptake of [¹⁴C]glycine was also measured to determine whether glycine might be a transmitter of the DNLL pathways to the IC. Lesions of the left DNLL failed to alter the Ca²⁺-dependent release or the uptake of [¹⁴C]glycine, suggesting that DNLL neurons are unlikely to use this compound as a transmitter.