Electron Paramagnetic Resonance Studies of Succinate:Ubiquinone Oxidoreductase from Paracoccus denitrificans

Electron paramagnetic resonance (EPR) studies of succinate:ubiquinone oxidoreductase (SQR) from Paracoccus denitrificans have been undertaken in the purified and membrane-bound states. Spectroscopic “signatures” accounting for the three iron-sulfur clusters (2Fe-2S, 3Fe-4S, and 4Fe-4S), cytochromeb, flavin, and protein-bound ubisemiquinone radicals have been obtained in air-oxidized, succinate-reduced, and dithionite-reduced preparations at 4–10 K. Spectra obtained at 170 K in the presence of excess succinate showed a signal typical of that of a flavin radical, but superimposed with another signal. The superimposed signal originated from two bound ubisemiquinones, as shown by spectral simulations. Power saturation measurements performed on the air-oxidized enzyme provided evidence for a weak magnetic dipolar interaction operating between the oxidized 3Fe-4S cluster and the oxidized cytochrome b. Power saturation experiments performed on the succinate- and dithionite-reduced forms of the enzyme demonstrated that the 4Fe-4S cluster is coupled weakly to both the 2Fe-2S and the 3Fe-4S clusters. Quantitative interpretation of these power saturation experiments has been achieved through redox calculations. They revealed that a spin-spin interaction between the reduced 3Fe-4S cluster and the cytochrome b (oxidized) may also exist. These findings form the first direct EPR evidence for a close proximity (≤2 nm) of the high potential 3Fe-4S cluster, situated in the succinate dehydrogenase part of the enzyme, and the low potential, low spin b-heme in the membrane anchor of the enzyme

Changes in the geographical distribution and abundance of the tick Ixodes ricinus during the past 30 years in Sweden

<p>Abstract</p> <p>Background</p> <p><it>Ixodes ricinus </it>is the main vector in Europe of human-pathogenic Lyme borreliosis (LB) spirochaetes, the tick-borne encephalitis virus (TBEV) and other pathogens of humans and domesticated mammals. The results of a previous 1994 questionnaire, directed at people living in Central and North Sweden (Svealand and Norrland) and aiming to gather information about tick exposure for humans and domestic animals, suggested that <it>Ixodes ricinus </it>ticks had become more widespread in Central Sweden and the southern part of North Sweden from the early 1980s to the early 1990s. To investigate whether the expansion of the tick's northern geographical range and the increasing abundance of ticks in Sweden were still occurring, in 2009 we performed a follow-up survey 16 years after the initial study.</p> <p>Methods</p> <p>A questionnaire similar to the one used in the 1994 study was published in Swedish magazines aimed at dog owners, home owners, and hunters. The questionnaire was published together with a popular science article about the tick's biology and role as a pathogen vector in Sweden. The magazines were selected to get information from people familiar with ticks and who spend time in areas where ticks might be present.</p> <p>Results</p> <p>Analyses of data from both surveys revealed that during the near 30-year period from the early 1980s to 2008, <it>I. ricinus </it>has expanded its distribution range northwards. In the early 1990s ticks were found in new areas along the northern coastline of the Baltic Sea, while in the 2009 study, ticks were reported for the first time from many locations in North Sweden. This included locations as far north as 66°N and places in the interior part of North Sweden. During this 16-year period the tick's range in Sweden was estimated to have increased by 9.9%. Most of the range expansion occurred in North Sweden (north of 60°N) where the tick's coverage area doubled from 12.5% in the early 1990s to 26.8% in 2008. Moreover, according to the respondents, the abundance of ticks had increased markedly in LB- and TBE-endemic areas in South (Götaland) and Central Sweden.</p> <p>Conclusions</p> <p>The results suggest that <it>I. ricinus </it>has expanded its range in North Sweden and has become distinctly more abundant in Central and South Sweden during the last three decades. However, in the northern mountain region <it>I. ricinus </it>is still absent. The increased abundance of the tick can be explained by two main factors: First, the high availability of large numbers of important tick maintenance hosts, i.e., cervids, particularly roe deer (<it>Capreolus capreolus</it>) during the last three decades. Second, a warmer climate with milder winters and a prolonged growing season that permits greater survival and proliferation over a larger geographical area of both the tick itself and deer. High reproductive potential of roe deer, high tick infestation rate and the tendency of roe deer to disperse great distances may explain the range expansion of <it>I. ricinus </it>and particularly the appearance of new TBEV foci far away from old TBEV-endemic localities. The geographical presence of LB in Sweden corresponds to the distribution of <it>I. ricinus</it>. Thus, LB is now an emerging disease risk in many parts of North Sweden. Unless countermeasures are undertaken to keep the deer populations, particularly <it>C. capreolus </it>and <it>Dama dama</it>, at the relatively low levels that prevailed before the late 1970s - especially in and around urban areas where human population density is high - by e.g. reduced hunting of red fox (<it>Vulpes vulpes</it>) and lynx (<it>Lynx lynx</it>), the incidences of human LB and TBE are expected to continue to be high or even to increase in Sweden in coming decades.</p

Processing of Bacillus subtilis succinate dehydrogenase and cytochrome b-558 polypeptides

The DNA sequence of the Bacillus subtilis sdh operon coding for the two succinate dehydrogenase subunits and cytochrome b-558 (the membrane anchor protein) has recently been established. We have now determined the extent of N-terminal processing of each polypeptide by radiosequence analysis. At the same time, direct evidence for the correctness of the predicted reading frames has been obtained. The cytochrome showed a ragged N-terminus, with forms lacking one residue, and is inserted across the membrane without an N-terminal leader-peptide. Covalently bound flavin was not detectable in B. subtilis succinate dehydrogenase expressed in Escherichia coli despite normal N-terminal processing of the apoprotein. This provides an explanation to why the succinate dehydrogenase synthesized in E. coli is not functional and demonstrates that host-specific factors regulate the coenzyme attachment

Bacillus subtilis CtaA is a heme-containing membrane protein involved in heme A biosynthesis.

Heme A is a prosthetic group of many respiratory oxidases. It is synthesized from protoheme IX (heme B) seemingly with heme O as a stable intermediate. The Bacillus subtilis ctaA and ctaB genes are required for heme A and heme O synthesis, respectively (B. Svensson, M. Lübben, and L. Hederstedt, Mol. Microbiol. 10:193-201, 1993). Tentatively, CtaA is involved in the monooxygenation and oxidation of the methyl side group on porphyrin ring D in heme A synthesis from heme B. B. subtilis ctaA and ctaB on plasmids in both B. subtilis and Escherichia coli were found to result in a novel membrane-bound heme-containing protein with the characteristics of a low-spin b-type cytochrome. It can be reduced via the respiratory chain, and in the reduced state it shows light absorption maxima at 428, 528, and 558 nm and the alpha-band is split. Purified cytochrome isolated from both B. subtilis and E. coli membranes contained one polypeptide identified as CtaA by amino acid sequence analysis, about 0.2 mol of heme B per mol of polypeptide, and small amounts of heme A