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J Wilms

Publications and source records attributed to J Wilms.

15 recordsLinked to original sources
PubMed1995

Detection of Epstein-Barr virus DNA in anal scrapings from HIV-positive homosexual men.

Epstein-Barr virus (EBV) can infect epithelial cells as well as B lymphocytes. Infection of the male and female genital tracts has recently been demonstrated, and it has been suggested that the virus may be sexually transmissible. In our study we investigated whether EBV can be found in the anal region of sexually active homosexual men. Anal scrapings from HIV-positive homosexual men and a heterosexual control population were investigated using the polymerase chain reaction (PCR) to screen for EBV DNA. EBV DNA was detected in 8 of 27 anal samples (29.6%) from the homosexual men and 3 of 34 samples (8.8%) from the heterosexual men. Our study shows that, like the genital tract, the anal region can harbour EBV subclinically. This finding suggests that the anal region may be a reservoir for EBV and that sexual transmission of this virus may be possible.

Anal Canal↗
PubMed1981

The effect of pH and ionic strength on the pre-steady-state reaction of cytochrome c and cytochrome aa3.

(1) In the pH range between 5.0 and 8.0, the rate constants for the reaction of ferrocytochrome c with both the high- and low-affinity sites on the cytochrome aa3 increased by a factor of approx. 2 per pH unit. (2) The pre-steady-state reaction between ferrocytochrome c and cytochrome aa3 did nt cause a change in the pH of an unbuffered medium. Furthermore, it was found that this reaction and the steady-state reaction are equally fast in H2O and 2H2O. From these results it was concluded that no protons are directly involved in a rate-determining reaction step. (3) Arrhenius plots show that the reaction between ferrocytochrome c and cytochrome aa3 requires a higher enthalpy of activation at temperatures below 20 degrees C (15--16 kcal/mol) as compared to that at higher temperature (9 kcal/mol). We found no effect of ionic strength on the activation enthalpy of the pre-steady-state reaction, nor on that of the steady-state reaction. This suggests that ionic strength does not change the character of these reactions, but merely affects the electrostatic interaction between both cytochromes.

Animals↗
PubMed1981

The reaction between cytochrome c1 and cytochrome c.

The kinetics of electron transfer between the isolated enzymes of cytochrome c1 and cytochrome c have been investigated using the stopped-flow technique. The reaction between ferrocytochrome c1 and ferricytochrome c is fast; the second-order rate constant (k1) is 3.0 . 10(7) M-1 . s-1 at low ionic strength (I = 223 mM, 10 degrees C). The value of this rate constant decreases to 1.8 . 10(5) M-1 . s-1 upon increasing the ionic strength to 1.13 M. The ionic strength dependence of the electron transfer between cytochrome c1 and cytochrome c implies the involvement of electrostatic interactions in the reaction between both cytochromes. In addition to a general influence of ionic strength, specific anion effects are found for phosphate, chloride and morpholinosulphonate. These anions appear to inhibit the reaction between cytochrome c1 and cytochrome c by binding of these anions to the cytochrome c molecule. Such a phenomenon is not observed for cacodylate. At an ionic strength of 1.02 M, the second-order rate constants for the reaction between ferrocytochrome c1 and ferricytochrome c and the reverse reaction are k1 = 2.4 . 10(5) M-1 . s-1 and k-1 = 3.3 . 10(5) M-1 . s-1, respectively (450 mM potassium phosphate, pH 7.0, 1% Tween 20, 10 degrees C). The 'equilibrium' constant calculated from the rate constants (0.73) is equal to the constant determined from equilibrium studies. Moreover, it is shown that at this ionic strength, the concentrations of intermediary complexes are very low and that the value of the equilibrium constant is independent of ionic strength. These data can be fitted into the following simple reaction scheme: cytochrome c2+1 + cytochrome c3+ in equilibrium or formed from cytochrome c3+1 + cytochrome c2+.

Animals↗
PubMed1981

Ionic strength effects on cytochrome aa3 kinetics.

1. The occurrence of an optimal ionic strength for the steady-state activity of isolated cytochrome aa3 can be attributed to two opposite effects: upon lowering of the ionic strength the affinity between cytochrome c and cytochrome aa3 increases, whereas in the lower ionic strength region the formation of a less active cytochrome c-aa3 complex limits the ferrocytochrome c association to the low affinity site. 2. At low ionic strength, the reduction of cytochrome c-aa3 complex by ferrocytochrome c1 proceeds via non-complex-bound cytochrome c. Under these conditions the positively charged cytochrome c provides the electron transfer between the negatively charged cytochromes c1 and aa3. 3. Polylysine is found to stimulate the release of tightly bound cytochrome c from the cytochrome c-aa3 complex. This property points to the existence of negative cooperativity between the two binding sites. We suggest that the stimulation is not restricted to polylysine, but also occurs with cytochrome c. 4. Dissociation rates of both high and low affinity sites on cytochrome aa3 were determined indirectly. The dissociation constants, calculated on the basis of pre-steady-state reaction rates at an ionic strength of 8.8 mM, were estimated to be 0.6 nM and 20 microM for the high and low affinity site, respectively.

Animals↗
PubMed1981

Properties and hepatic metabolism of non-transferrin-bound iron.

These experiments have examined the physical properties and the hepatic uptake of non-transferrin-bound iron, a fraction found in the serum of untreated primary hemochromatosis patients. Ultrafiltration, polyacrylamide gel electrophoresis, and TLC have demonstrated the non-transferrin-bound iron complex to have a molecular weight less than 1000 and to be distinct from free iron and amino acid iron complexes. Hepatic iron uptake from serum containing non-transferrin-bound iron was significantly higher than from the same serum sample from which the abnormal iron had been removed. Non-transferrin-bound iron entering the liver was localized to the lysosomal fraction in the first hour and was subsequently incompletely incorporated into ferritin. The results demonstrated an abnormal uptake of this iron fraction by the liver and suggest that the abnormal serum fraction may contribute to tissue damage by entering the lysosomes.

Animals↗
PubMed1980

The effect of pH and ionic strength on the steady-state activity of isolated cytochrome C oxidase.

1. The turnover number and apparent Km of isolated beef-heart cytochrome c oxidase were found to increase continuously when the pH was lowered from 8.6 to 4.6 (turnover number 32-630 s-1). In this pH range neither irreversible denaturation of the enzyme nor an optimum for the turnover number was observed. 2. The turnover number of cytochrome c oxidase was found to be independent of ionic strength. It was concluded that the dependence of the activity of cytochrome c oxidase on ionic strength is caused by a change in the value of Km for cytochrome c. 3. The pH dependence of the turnover number of cytochrome c oxidase can be described by a simple model in which at least three sites on the complex of cytochrome c oxidase with cytochrome c (pKa 8.0, 6.5 and 4.8) can take up a proton.

Cytochrome c Group↗
PubMed1980

The pre-steady state reaction of ferrocytochrome c with the cytochrome c-cytochrome aa3 complex.

1. Using stopped-flow technique we have investigated the electron transfer form cytochrome c to cytochrome aa3 and to the (porphyrin) cytochrome c-cytochrome aa3 complex. 2. In a low ionic strength medium, the pre-steady state reaction occurs in a biphasic way with rate constants of at least 2.10(8) M-1.s-1 and about 10(7) M-1.S-1 (I=8.8 mM, pH 7.0, 10 degrees C), respectively. 3. A comparison of the rate constants, determined in the presence of an excess of cytochrome c with those found in the presence of an excess of cytochrome aa3 reveals the existence of two slower reacting sites on the functional unit (2 hemes and 2 coppers) of cytochrome aa3. On basis of these results we discuss various models. If no site-site interactions are assumed (non-cooperative model) cytochrome aa3 has 2 high and 2 low affinity sites available for the reaction with ferrocytochrome c. If negative cooperativity occurs, cytochrome aa3 has 2 high affinity sites which change into 2 low affinity sites upon binding of one cytochrome c molecule. The latter model is favoured.

Animals↗
PubMed1980

The pre-steady state and steady-state kinetics of the ATPase activity of mitochondrial F1.

1. The lag time before maximum velocity of ATP hydrolysis is reached upon mixing ATP with F1 is much greater than can be explained by a simple Michaelis-Menten mechanism, and must be due to an activation reaction. The lag time is dependent on the concentration of MgATP (half-maximal at 30 microM) and is equal to 30 ms at infinite MgATP concentration. The initial rate of hydrolysis by nucleotide-depleted F1 is much greater than with normal F1. It is tentatively suggested that the activation reaction with normal preparations is due to replacement of firmly bound ADP by MaATP. 2. After the initial time lag, the reaction follows very closely first-order kinetics provided that the concentration of MgATP is much less than the Km and the reaction is completed within 2 s. This is not expected if the dissociation constant of the enzyme-MgADP complex, an intermediate in the enzymic reaction, is much lower than the Km as has been reported in the literature. The value of V/Km, calculated from the exponential decay, is very close to that calculated from independent measurements of V and Km. 3. The low values for Ki(ADP) reported in the literature were found to be due to a slow (in the order of seconds) formation of an inhibited MgADP-enzyme complex. Dissipation of this inhibited complex by ATP requires seconds. The dissociation constant of the MgADP-enzyme complex that is an intermediate in the enzyme reaction was found to be 150 microM. 4. ADP but not ATP becomes firmly bound to nucleotide-depleted F1 in the absence of Mg2+.

Adenosine Diphosphate↗
PubMed1980

Reactions of mercaptans with cytochrome c oxidase and cytochrome c.

1. The steady-state oxidation of ferrocytochrome c by dioxygen catalyzed by cytochrome c oxidase, is inhibited non-competitively towards cytochrome c by methanethiol, ethanethiol, 1-propanethiol and 1-butanethiol with Ki values of 4.5, 91, 200 and 330 microM, respectively. 2. The inhibition constant Ki of ethanethiol is found to be constant between pH 5 and 8, which suggests that only the neutral form of the thiol inhibits the enzyme. 3. The absorption spectrum of oxidized cytochrome c oxidase in the Soret region shows rapid absorbance changes upon addition of ethanethiol to the enzyme. This process is followed by a very slow reduction of the enzyme. The fast reaction, which represents a binding reaction of ethanethiol to cytochrome c oxidase, has a k1 of 33 M-1 . s-1 and a dissociation constant Kd of 3.9 mM. 4. Ethanethiol induces fast spectral changes in the absorption spectrum of cytochrome c, which are followed by a very slow reduction of the heme. The rate constant for the fast ethanethiol reaction representing a bimolecular binding step is 50 M-1 . s-1 and the dissociation constant is about 2 mM. Addition of up to 25 mM ethanethiol to ferrocytochrome c does not cause spectral changes. 5. EPR (electron paramagnetic resonance) spectra of cytochrome c oxidase, incubated with methanethiol or ethanethiol in the presence of cytochrome c and ascorbate, show the formation of low-spin cytochrome alpha 3-mercaptide compounds with g values of 2.39, 2.23, 1.93 and of 2.43, 2.24, 1.91, respectively.

Animals↗
PubMed1979

Inhibition of mitochondrial electron transfer in rats by ethanethiol and methanethiol.

1. We have investigated the effects of ethanethiol, methanethiol and dimethyl sulphide on some metabolic processes of isolated rat hepatocytes, isolated mitochondria from liver and brain and ox-heart submitochondrial particles. 2. Ethanethiol, but not dimethyl sulphide, inhibited both gluconeogenesis and ureogenesis from various substrates in rat hepatocytes, depressed cellular ATP content and caused an increased reduction of the mitochondria. 3. Ethanethiol inhibited respiration in isolated rat-liver mitochondria with several substrates, both in the presence of ADP and phosphate or in the presence of an uncoupling agent. Ethanethiol also inhibited respiration in isolated rat-brain mitochondria. Dimethyl sulphide was much less effective in inhibiting mitochondrial respiration. 4. In ox-heart submitochondrial particles ethanethiol inhibited electron transfer between cytochrome c and oxygen. 5. Purified cytochrome c oxidase was inhibited by ethanethiol in a non-competitive manner. 6. Methanethiol inhibited cytochrome c oxidase and was an effective inhibitor of mitochondrial electron transfer, both in liver and brain. 7. The difference in inhibitory properties between ethanethiol, methanethiol and dimethyl sulphide observed in our experiments coincides with the difference in potency to elicit coma in rats. We suggest that inhibition of mitochondrial electron transfer by mercaptans may be relevant to the mechanism by which energy production in brain is depressed during hepatic coma.

Animals↗