Uptake Apparatus Research Articles

Natural genetic exchange between Haemophilus and Neisseria: intergeneric transfer of chromosomal genes between major human pathogens.

Members of the bacterial families Haemophilus and Neisseria, important human pathogens that commonly colonize the nasopharynx, are naturally competent for DNA uptake from their environment. In each genus this process is discriminant in favor of its own and against foreign DNA through sequence specificity of DNA receptors. The Haemophilus DNA uptake apparatus binds a 29-bp oligonucleotide domain containing a highly conserved 9-bp core sequence, whereas the neisserial apparatus binds a 10-bp motif. Each motif ("uptake sequence", US) is highly over-represented in the chromosome of the corresponding genus, particularly concentrated with core sequences in inverted pairs forming gene terminators. Two Haemophilus core USs were unexpectedly found forming the terminator of sodC in Neisseria meningitidis (meningococcus), and sequence analysis strongly suggests that this virulence gene, located next to IS1106, arose through horizontal transfer from Haemophilus. By using USs as search strings in a computer-based analysis of genome sequence, it was established that while USs of the "wrong" genus do not occur commonly in Neisseria or Haemophilus, where they do they are highly likely to flag domains of chromosomal DNA that have been transferred from Haemophilus. Three independent domains of Haemophilus-like DNA were found in the meningococcal chromosome, associated respectively with the virulence gene sodC, the bio gene cluster, and an unidentified orf. This report identifies intergenerically transferred DNA and its source in bacteria, and further identifies transformation with heterologous chromosomal DNA as a way of establishing potentially important chromosomal mosaicism in these pathogenic bacteria.

Measurement and interpretation of equilibrium organic vapor sorption in polyurethane membranes

The equilibrium sorption of benzene, tetrachloroethylene, and hexane vapor in four commercially available polyurethanes was measured as a function of temperature (30–60°C) and organic activity with a McBain vapor uptake apparatus. The four polyurethanes were composed of the same polyether soft segments, but contained different amounts of phase-separated, hydrogen-bonded hard segment domains. For all three organic penetrants, polymer swelling was inversely proportional to the polyurethane's hard segment content. Vapor sorption data were modeled over the entire organic activity range for all four polyurethanes using the Flory–Rhener equation (the Flory–Huggins equation did not fit the experimental data at high organic vapor activities). The morphology of each of the polyurethanes was analyzed by FTIR (in terms of the hydrogen bonding index, HBI), elemental analysis (to calculate the total volume of hard segments in a polyurethane), and small-angle X-ray scattering (to determine the long spacing between hard segment domains). The Flory–Huggins interaction parameters, χ, in the Flory–Rhener equation for the three organic vapors were found to increase linearly with HBI. The M c parameter in the Flory–Rhener equation for each polymer, defined as the effective molecular weight of soft segments between hard domains, was correlated empirically with a combination of the HBI, polymer hard segment content, and soft segment long spacing.

Is there a relationship between baseline and treatment-associated changes in [3H]-IMI platelet binding and clinical response in major depression?

A peripheral model for the central 5-HT neuron is the characterization of platelet imipramine binding. We studied an outpatient major depressive cohort who fulfilled Research Diagnostic Criteria for agitation. After a 1-week placebo lead-in, subjects were blindly randomized to either imipramine (IMI) or fluoxetine (FLU) during an 8-week, double-blind study period. Thirty-three subjects (15 IMI, 18 FLU) provided both baseline and endpoint samples for the platelet [3H]-IMI assay. Depression efficacy was comparable across the two treatments, whereas FLU was significantly more effective in reducing secondary anxiolysis (p = .023). Discontinuations due to an adverse event were significantly more frequent with IMI than FLU (p < .01). Baseline affinity (KD) was mildly predictive of change in the HAMD (r = -.22; p = .07). Whereas baseline to endpoint density (Bmax) changes (delta) were similar for IMI (183 +/- 329 fmol/mg) and FLU (196 +/- 402 fmol/mg), a statistically significant treatment difference in delta KD emerged (IMI -0.005 +/- 0.010 pmol/ml versus FLU 0.008 +/- 0.013 at p = 004). Moreover, the changes in KD and HAMD17 trended to a positive correlation among only the FLU-treated subjects (4 = 0.406, p = .095). The clinical effects of 5-HT-based selective antidepressant may be reflected by dynamic changes in the platelet 5-HT uptake apparatus. These data suggest that the baseline confirmational status of the [3H]-IMI:5-HT transporter may reflect a "capacity" for a treatment response.

Prepilin-like molecules in type 4 pilus biogenesis: minor subunits, chaperones or mediators of organelle translocation?

identity across diverse species boundaries, have been proposed to be part of similar apparatuses for extracellular protein localization. These include genes encoding type 4 prepilin-like polypeptides, peptidases responsible for amino-terminal processing of the prepilin homologues, proteins peripherally associated with the cytoplasmic membrane with highly conserved nucleosidetriphosphate-binding (NTP-binding) sites, and outer-membrane proteins. Homologues corresponding to one or more of these families have been implicated in the extracellular localization of toxins and hydrolases by Gram-negative bacteria. Collectively, this has been termed the type II system: the two-step protein-export system or the general secretion pathway (GSP)‘. Related components are also essential for filamentous phage morphogenesis! and for the proper construction, localization and function of a DNAbinding and -uptake apparatus associated with competence for natural transformation in Bacillus subtilis’ and Haemophilus influenzaes. For all of these systems, the goal of research has been to understand how a structure analogous to type 4 pili, or events related to pilus biogenesis, might link these varied processes. I would like to suggest that perhaps we should look at the situation from the other point of view, and examine what studies of the GSP might reveal about type 4 pilus biogenesis.

ComK encodes the competence transcription factor, the key regulatory protein for competence development in Bacillus subtilis

comK is a positive autoregulatory gene occupying a central position in the competence-signal-transduction network. All regulatory routes identified in this network converge at the level of comK expression. The ComK protein is required for the transcriptional induction of comK and the late competence genes, which specify morphogenetic and structural proteins necessary for construction of the DNA-binding and uptake apparatus. In this report we demonstrate that ComK specifically binds to DNA fragments containing promoter and upstream sequences of the genes it affects (comC, comE, comF, comG and comK). Using portions of the region upstream of comC we show that the ComK-binding sequences are essential for the expression of competence. Moreover, we demonstrate that the presence of ComK stimulates the expression of comF-lacZ and comG-lacZ translational fusions in vivo in Escherichia coli. These results indicate that the gene product of comK is identical to the previously inferred competence transcription factor (CTF).

Antiproliferative properties of polyamine analogues: a structure-activity study.

A basis set of polyamine analogues was designed and synthesized. These compounds were used to initiate a systematic investigation of the role of chain length, terminal nitrogen alkyl group size, and symmetry of the methylene backbone in the antineoplastic properties of polyamine analogues. New synthetic methods predicated on our earlier polyamine fragment synthesis are described for accessing the tetraamines of interest. An unsymmetrically substituted diamine reagent, N-(tert-butoxycarbonyl)-N,N'-bis(mesitylenesulfonyl)-1,4-diaminobu tane, was developed for entry into unsymmetrical tetraamines. All of the tetraamines synthesized were first evaluated in a murine leukemia L1210 cell IC50 assay at 48 and 96 h. In an attempt to correlate this behavior with some aspect of polyamine metabolism, each compound was tested for its ability to compete with spermidine for the polyamine uptake apparatus, its impact on the polyamine biosynthetic enzymes ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (AdoMetDC), and its effect on the polyamine-catabolizing enzyme spermidine/spermine N1-acetyltransferase (SSAT) and on polyamine pools. While there was no obvious correlation between the 48 and 96 h IC50's and the impact of the analogues on polyamine metabolism, there were other structure-activity relationships. Correlations were observed to exist between chain length and IC50's and between terminal alkyl substituents and impact on Ki, ODC, and AdoMetDC. Also, preliminary studies suggest a relationship may exist between the 48 and 96 h IC50 activities and the analogue's chronic toxicity in vivo. Finally, when the overall length of the polyamine backbone was held constant, the symmetry of the methylene chains of the polyamine fragments was shown to be unimportant to the compound's activity.

Thermal and photooxidative studies of polyethylene

AbstractThe technique of oxygen uptake measurement for assessing the photo‐ and thermal stability of polyethylene is reviewed. An oxygen uptake apparatus is described which was used to determine the thermooxidative stabilities of nascent, high‐density polyethylenes produced by Ziegler‐Natta, and silica‐supported “silyl chromate” and “chromocene” catalyst systems. The oxygen uptake results are compared with those obtained from the thermochemical technique of differential scanning calorimetry.

Hexahydropyrimidines as masked spermidine vectors in drug delivery

N-(4-Aminobutyl)hexahydropyrimidine and N-(3-aminopropyl)hexahydropyrimidine are shown to compete with spermidine for uptake by L1210 cells. This observation is in keeping with the idea that spermidine may adopt a hydrogen-bonded cyclic structure in the course of transport. Furthermore, the differences in the ability of spermidine, homospermidine, and norspermidine to utilize the spermidine uptake apparatus is analyzed in terms of the charge on the triamines, as calculated from p K a data.

Identification of mitochondrial proteins and some of their precursors in two-dimensional electrophoretic maps of human cells.

A set of at least 30 proteins disappears from the two-dimensional electrophoretic pattern of human lymphoid cells treated with various antimitochondrial agents. This set is similar to the set of proteins found in isolated mitochondria (except for the presence of action in the latter group), indicating that the inhibitor effect stops production of a majority of mature mitochondrial proteins. Several proteins having the characteristics of precursors to the major cytoplasmically synthesized mitochondrial proteins can be observed in cells during fast-pulse experiments and in a reticulocyte lysate system fed with total lymphoid cell RNA. In the three major instances of mitochondrial precursor--product processing, the removed peptide is quite basic in each case, suggesting that a lysine- or arginine-rich terminal sequence may be necessary for initial recognition by the mitochondrial protein uptake apparatus. The inhibitor effect allows easy identification of a large set of mitochondrial proteins in two-dimensional maps of various cells, thereby specifying a particularly tractable and functionally distinctive subset of the cellular proteins. The nature and wide scope of the effect support the concept of energy-dependent "vectorial processing" [Schatz, G. (1979) FEBS Lett. 103, 203--211] and indicate that such a mechanism is generally applicable to the major class of cytoplasmically synthesized mitochondrial proteins in mammalian cells.