Martin N. Hughes

Toxicity and accumulation of thallium in bacteria and yeast

Thallium sulphate inhibited microbial growth, withBacillus megaterium KM, more sensitive to the metal thanSaccharomyces cerevisiae andEscherichia coli. Inhibition ofB. megaterium KM andS. cerevisiae, but not ofE. coli, was alleviated by increasing the potassium concentration of the medium; inhibition of respiration ofS. cerevisiae, but not ofE. coli, was similarly alleviated. Thallium was rapidly bound, presumably to cell surfaces, byS. cerevisiae andE. coli, and was progressively accumulated by energy-dependent transport systems (probably concerned primarily with potassium uptake) with both organisms. Thallium uptake kinetics suggested more than one transport system operated in yeast, possibly reflecting a multiplicity of potassium transport systems. ApparentKm andKi values for competitive inhibition of thallium uptake by potassium indicatedS. cerevisiae to have a higher affinity for thallium uptake than for potassium, whileE. coli had a transport system with a higher affinity for potassium than for thallium. The likely systems for thallium transport are discussed. A mutant ofE. coli with tenfold decreased sensitivity to thallium was isolated and apparently effected surface binding of thallium in amounts equivalent to the wild type organism, but showed no subsequent uptake and accumulation of the metal from buffer, even though it was able to accumulate potassium to normal intracellular concentrations during growth.

Nitrate, but not silver, ions induce spectral changes in Escherichia coli cytochrome d

The absorbance maximum (630 nm) of reduced cytochrome d in Escherichia coli membrane particles was diminished by 160 μM AgNO3 or NaNO3 and accompanied by the formation of a species with an absorption maximum at 640–645 nm. Nitrite, trioxodinitrate and nitric oxide elicited qualitatively similar, but faster, changes in the spectrum of cytochrome d, suggesting that formation of a nitrosyl complex may be involved in all cases. In direct contrast to an earlier report, silver ions (160 υM) were without effect on the α‐bands of reduced cytochromes d, b or a 1.

Filamentous growth of Escherichia coli K12 elicited by dimeric, mixed-valence complexes of ruthenium.

Dimeric, mixed-valence [(Ru(II), Ru(III)] compounds of ruthenium caused filament formation in growing cultures of Escherichia coli K12. Three compounds with the general formula Ru2(NH3)6X5 · H2O (where X is a halide) were tested; in order of decreasing effectiveness (and with the concentration giving maximum effect), these were the bromo (10-5M), chloro (10-4 to 10-5M), and iodo (10-3 to 10-4M) analogues. Filamentation elicited by the bromo and chloro compounds was spontaneously reversible after 3–4 h, and tentatively attributed to oxidation of the active mixed-valence form to inactive Ru(III) complexes. Several compounds known to accelerate division of filaments formed under other conditions were ineffective in reversing the filamentation, but the presence of 0,43 M-dimethylsulphoxide totally inhibited filamentation caused by the bromo or chloro compounds and by cis-Pt(NH3)2Cl2 (cisplatin), an established filamenting and antitumour agent. The ruthenium complexes bound to mammalian DNA, but were without effect on the UV spectrum or cellular content of DNA in E. coli, despite showing marked mutagenic activity in reverse mutation tests with Salmonella typhimurium. Cells remained sensitive to inhibition of division by the ruthenium complexes until immediately prior to the division event. Possibilities for the (probably complex) mode of action and the potential of related compounds for therapeutic use are discussed.

The toxicity of thallium(I) to cardiac and skeletal muscle.

The effects of (a) partial or complete replacement of K+ by Tl+ in saline perfusing isolated rat heart and diaphragm preparations and (b) pulse injections of high concentrations of Tl+ or K+, have been studied. The immediate effect of Tl+ resembles that of higher concentrations of K+ and may reflect its more rapid penetration into the tissue. Tl+ appears to replace K+ on a 1:1 basis to an extent dependent upon the relative abundance of the two cations in the perfusion solution. However, analysis of diaphragm preparations after perfusion with salines containing increasing Tl+ but constant [K+ + Tl+] showed a related and progressive increase in total cation content. This effect, which was not seen in the presence of constant high (normal) K+ concentrations, may reflect an increase of the intracellular space brought about by the thallium. Functional effects of Tl+ were (a) preferential block of the phrenic nerve or neuromuscular junction over the muscle fibre and (b) transient but marked acceleration of cardiac frequency following pulse injections, which may be of value in analysing the pacemaker mechanism of the heart. In both tissues Tl+ is eventually toxic and probably irreversibly so.

Resonance Raman spectroscopy of an oxygenated intermediate species of cytochrome oxidase d from Escherichia coli

Resonance Raman spectroscopy (excitation at 647.1 nm) of solubilized and aerated cytochrome oxidase d from Escherichia coli K12 has shown absorptions (1078–1105 cm−1) attributed to the oxygen—oxygen stretching frequency of the oxidase—oxygen adduct. These findings support the hypothesis that the 650–652 nm chromophore of cytochrome d is an oxygenated or ’oxy’ intermediate species and not the fully oxidized enzyme.

Metal complexes of hydroxylamine

Abstract The following complexes with hydroxylamine or deprotonated hydroxylamine as a ligand have been prepared and characterised: [NiL 6 ]X 2 , X = Cl, Br, ClO 4 ; NiL 4 X 2 , X = Cl, Br, NO 3 , ClO 4 ; NiL 3 Cl 2 ; NiL 2 X 2 , X = Cl, Br; NiLX 2 , X = Cl, NO 3 ; Ni 2 L 6 (L−H)X 3 , X = Cl, Br; NiL 4 (L−H) 2 ; ZnL 2 X 2 , X = Cl, Br, 1 2 SO 4 ; Zn 2 L 3 (L−H)X 3 , X = Cl, Br; CdL 2 X 2 , X = Cl, Br; CoL 2 X 2 , X = Cl, 1 2 SO 4 . A few complexes with O-methylhydroxylamine have been obtained, but there is no evidence for complex formation in solution between metal halides and N-methylhydroxylamine. All the nickel compounds are six coordinate, the ligand is nitrogen bonded to the metal and present as NH 2 OH, with Δ o (in [NiL 6 ]X 2 ) = 12.6 kK. The product from Ni(OH) 2 and NH + 3 OHCl − has the hydroxylamine present in a different form, probably the N-oxide, as evidenced by infrared spectroscopy. Two forms of ZnL 2 X 2 have been prepared, the course of the reaction being solvent dependent. Cd(II) and Co(II) 1:2 complexes contain the ligand in the N-oxide form, M(ONH 3 )X 2 .

The far infrared spectra of some complexes of palladium (II) with thiazoles

Abstract The infrared spectra of six pairs of palladiuim(II) chloride and bromide complexes are reported. The bromo complexes with non 2-substituted ligands appear to show high metal-bromine frequencies.

Thallium activation and inhibition of yeast aldehyde dehydrogenase

Volume 59, number 1 FEBS LETTERS November 1975 THALLIUM ACTIVATION AND INHIBITION OF YEAST ALDEHYDE DEHYDROGENASE K. BOSTIAN and G. F. BETTS Department of Plant Biology and Microbiology, Queen Mary College, Mile End Road, London, E. 1. England W. K. MAN and M. N. HUGHES Department of Chemistry, Queen Elizabeth College, Campden Hill Road, London, I¢. 8. England Received 1 September i975 1. Introduction T1 ÷ has been shown to replace the physiological ion K ÷ in the activation of several monovalent cation activated enzymes [1-6]. This provides the opportu- nity to employ T1 ÷ as a probe for K ÷ in n.m.r, and fluorescence studies [6-8]. Preliminary to such studies on yeast aldehyde dehydrogenase (EC 1.2.1.3.) T1 ÷ activation of this enzyme was investigated and compared to the previously demonstrated activation by K ÷ [9]. A general mechanism for this activation has been proposed whereby an activating monovalent cation induces and maintains an enzyme conformation suitable for catalytic activity [ 10]. 2. Materials and methods Partially purified aldehyde dehydrogenase was prepared from N.G. and S.F. yeast (British Fermenta- tion Products Ltd.) obtained from a local baker. The method was essentially that of Black [9] except for the inclusion of 1.2 X 10 -3 M phenylmethylsulphonyl fluoride (PMSF) throughout the procedure. The buffer composition at the final stage contained 0.1 M K2HPO4, pH 7.8, 10 -3 M mercaptoethanol, and 1.2 × 10 -3 M PMSF. Enzyme activity was between 7-10 units/ml when assayed with optimal K ÷ (0.1 M) with a specific activity of 1. Samples were stored frozen at -18 ° C, and thawed out immediately prior to use. Enzyme was equilibrated with 0.1 M Tris-NO3 pH 8.0, 10 -3 M mercaptoethanol and activating salt where indicated, by passage through a Sephadex G-25 column at 4°C. Because of the instability of the enzyme in the absence of an activating monovalent cation, the enzyme was stored on ice following chromatography and all assays in a given run were completed within 5 rain. Enzyme activity was measured by following the rate of reduction of NAD ÷ by observing the increase in absorbance at 340 nm in a Pye Unicam SP1800 spectrophotometer using 1 cm quartz cells. Reaction mixtures contained in a final volume of 2.5 ml: 2 X 10 -3 M acetaldehyde, 5 X 10 -4 M NAD ÷, 0.1 M Tris-NO3 buffer, pH 8.0,

Thallium(I) derivatives of some aminoacids and other acidic ligands. Studies in solution and the solid state

Abstract Thallium(I) derivatives of a number of acidic ligands, including some aminoacids, carboxylic acids, phenols and thiols, have been prepared and characterised by mass-, U.V., and I.R. spectroscopy. Interaction between thallium(I) and certain of these ligands in aqueous solution has been studied by U.V and fluorescence spectroscopy, and formation constants measured in some cases. It is concluded that neither property is of particular utility in characterising the interaction of thalium(I) with biological macromolecules.

The vibrational spectrum of anhydrous and hydrated sodium hyponitfite

Abstract The Raman and i.r. spectra of anhydrous and hydrated sodium hyponitrite have been recorded and differences in1the hyponitrite frequencies have been observed. The assignment of mode ν 3 is confirmed at 697 cm −1 .