Respiratory components in acidophilic bacteria that respire on iron

Abstract

Microorganisms capable of aerobic respiration on ferrous ions are spread throughout eubacterial and archaebacterial phyla. Phylogenetically distinct organisms were shown to express spectrally distinct redox‐active biomolecules during autotrophic growth on soluble iron. A new iron‐oxidizing eubacterium, designated as strain Funis, was investigated. Strain Funis was judged to be different from other known iron‐oxidizing bacteria on the bases of comparative lipid analyses, 16S rRNA sequence analyses, and cytochrome composition studies. When grown autotrophically on ferrous ions, Funis produced conspicuous levels of a novel acid‐stable, acid‐soluble yellow cytochrome with a distinctive absorbance peak at 579 nm in the reduced state.

Stopped‐flow spectrophotometric kinetic studies were conducted on respiratory chain components isolated from cell‐free extracts of Thiobacillus ferrooxidans. Experimental results were consistent with a model where the primary oxidant of ferrous ions is a highly aggregated c‐type cytochrome that then reduces the periplasmic rusticyanin. The Fe(II)‐dependent, cytochrome c‐catalyzed reduction of the rusticyanin possessed three kinetic properties in common with corresponding intact cells that respire on iron: the same anion specificity, a similar dependence of the rate on the concentration of ferrous ions, and similar rates at saturating concentrations of ferrous ions     

Acetylene reduction by nitrogen-fixing blue-green algae

Summary Known nitrogen-fixing species of blue-green algae are capable of reducing acetylene to ethylene, but acetylene is not reduced by Anacystis nidulans, which does not fix nitrogen. Cycad root nodules which contain blue-green algae as endophytes reduce acetylene. Acetylene reduction is inhibited by carbon monoxide. Nitrate or ammonium-nitrogen has no immediate effect on algae reducing acetylene, but algae grown on nitrate-nitrogen gradually lose their capacity to reduce acetylene. Nitrate-nitrogen also inhibits heterocyst formation in these algae and there is a fairly direct correlation between the abundance of heterocysts in a particular sample and its capacity to reduce acetylene. Aphanizomenon flosaquae reduces acetylene and fixes nitrogen in unialgal culture and there is strong presumptive evidence that these reductions are carried out by the alga rather than by associated bacteria. The molar ratios of ethylene: ammonia produced vary within the range 1.4–1.8.     

Analysis of fidelity mechanisms with eukaryotic DNA replication and repair proteins

We are investigating the mechanisms for producing or avoiding errors during DNA synthesis catalyzed by DNA replication and repair proteins purified from eukaryotic sources. Using assays that monitor the fidelity of a single round of DNA synthesis in vitro, we have defined the error frequency and mutational specificity of the four classes of animal cell DNA polymerases (alpha, beta, delta, gamma), and the fidelity of an SV40 origin-dependent DNA replication complex in extracts of HeLa cells.     

Fluorometric, Chromatographic, and Spectronic Evidence for the Non-indolic Nature of Citrus Auxin

Evidence for the non-indolic nature of the new citrus auxinis presented on the basis of fluorometric properties, thin-layerchromatography, Ehrlich's colour reaction, paper electrophoresis,and the infra-red spectra determinations. Citrus auxin had alower Rf in TLC than IAA, did not give the typical indole reactionwith Ehrlich's reagent, and behaved differently in electrophoresis.The infra-red spectra also provided preliminary informationconcerning chemical structure. The hypothesis that indolic compoundsconstitute the only natural auxins in higher plants should berevised in view of this evidence that a non-indole auxin existsin higher plants.     

Development of High-Frequency Delivery System for Transposon Tn919 in Lactic Streptococci: Random Insertion in Streptococcus lactis subsp. diacetylactis 18-16

The conjugative transposon Tn919, originally isolated in Streptococcus sanguis FC1, is capable of low-frequency transfer (10⁻⁷ and 10⁻⁸ per recipient) on membrane filters to a wide number of streptococcal recipients including the industrially important lactic streptococci. The introduction of pMG600 (Lac⁺ Lax⁻; a lactose plasmid capable of conjugative transfer at high frequencies and which, in certain hosts, confers an unusual clumping phenotype) into a Streptococcus lactis CH919 donor, generating S. lactis CH001, resulted in a significant improvement in the transfer frequency of Tn919 to S. lactis CK50 (1.25 × 10⁻⁴ per recipient). In addition, these matings could be performed on agar surfaces, allowing the recovery of a greater number of recipients than with filter matings. Tn919 also transferred at high frequency to S. lactis subsp. diacetylactis 18-16S but not to Streptococcus cremoris strains. Insertion in 18-16S transconjugants generated from filter matings with an S. lactis CH919 donor was random, occurring at different sites on the chromosome and also in plasmid DNA. Thus, the conditions necessary for the practical exploitation of Tn919 in the targeting and cloning of genes from a member of the lactic streptococci, namely, high-frequency delivery and random insertion in host DNA, were achieved.     

Major histocompatibility (B) complex and sex effects on the phytohaemagglutinin wattle response

The influence of the major histocompatibility (B) complex and sex on the phytohaemagglutinin (PHA) wattle response was studied in 136 segregants (B2/B2, B2/B5 and B5/B5) of a fourth generation cross between inbred lines 6(1) and 15(1). At 6 weeks of age, chickens were injected with 100 micrograms purified PHA-P. Wattle thickness measurements were taken 4, 24, 48, 72 and 96 h after injection. Analysis of variance showed that 4 h after injection, males had a significantly higher response than females but the sex-genotype interaction was also significant. Females had higher responses than males 24 and 48 h after injection as a consequence of more rapid development and earlier resolution of the reaction in males. B2/B2 chickens had significantly lower responses than B5/B5 chickens 72 and 96 h after injection, signifying a faster late resolution phase in the B2/B2 genotype. The developmental and early resolution phases of the PHA wattle response were influenced by sex while the late resolution phase was influenced by B genotype.     

Allometric Root/Shoot Relationships and Predicted Water Uptake for Desert Succulents

Root morphology, shoot morphology, and water uptake for Agavedeserti and Ferocactus acanthodes of various sizes were studiedusing allometric relationships (y = ax^b) and a previously developedwater uptake model. Shoot surface area increased with shootvolume with an exponent b of 0.75 for both species. Root lengthand the ground area explored by the roots increased with shootsurface area with b's of 0.72 for A. deserti and 0.92 for F.acanthodes. Various sized individuals had about the same ratioof root length to explored ground area, with higher values occurringfor A. deserti. Predicted water uptake averaged over the exploredground area was approximately constant over a 10⁴-fold rangein shoot surface area, suggesting that shoot size confers nointraspecific competitive advantage for water uptake. For theroot lengths per explored ground area observed in the field,water uptake was predicted to be 85 per cent of maximal; wateruptake could be increased by the production of more rain roots.When differences in shoot volume were accounted for by allometry,small plants had relatively less shoot surface area and relativelymore root length per shoot volume than did large plants, whichmay be important for the water relations of seedling establishment. Agave deserti, Ferocactus acanthodes, allometry, desert succulents, root distribution, root length, seedling growth, seedling establishment, shoot surface area, shoot volume, water uptake     

Binding of ions and hydrophobic probes to alpha-lactalbumin and kappa-casein as determined by analytical affinity chromatography

The technique of analytical affinity chromatography was extended to characterize binding of ions and hydrophobic probes to proteins. Using the immobilized protein mode of chromatography, alpha-lactalbumin and kappa-casein were covalently attached to 200-nm-pore-diameter controlled-pore glass beads and accommodated for high-performance liquid chromatography. The existence of a high affinity binding site (Kdiss = 0.16 microM) (site I) for calcium ion in alpha-lactalbumin was confirmed by chromatography of [45Ca2+]. In addition, chromatography of the hydrophobic probes, 1-(phenylamino)-8-naphthalene-sulfonate (ANS)2 and 4,4'-bis[1-(phenylamino)-8-naphthalenesulfonate (bis-ANS) indicated that Ca2+ bound to a second site (presumably the zinc site or site II) with weaker affinity. Dissociation constants obtained for apo-alpha-lactalbumin were about 80 microM for ANS and 4.7 microM for bis-ANS in the absence of sodium ion. Addition of Ca2+ initially caused a reduction in surface hydrophobicity (lowered affinity for the probe dyes) followed by an increase at higher Ca2+ concentrations (greater than 0.5 mM), suggesting that occupancy of site II restores an apo-like conformation to the protein. Moreover, the effect of Zn2+ was similar to that observed in the higher Ca2+ concentration range, whereas Na+ apparently bound to site I. A calcium binding site of moderate affinity also exists in kappa-casein (Kdiss = 15.6 microM). A cluster of negative charges, probably including the orthophosphate group, most likely comprise this binding site. By preventing self-association, analytical affinity chromatography permits microscale characterization of ligand equilibria in proteins that are unaffected by protein-protein interactions.