Dissecting the Conformational Dynamics of the Bile Acid Transporter Homologue ASBTNM

Apical sodium-dependent bile acid transporter (ASBT) catalyses uphill transport of bile acids using the electrochemical gradient of Na⁺ as the driving force. The crystal structures of two bacterial homologues ASBT_NM and ASBT_Yf have previously been determined, with the former showing an inward-facing conformation, and the latter adopting an outward-facing conformation accomplished by the substitution of the critical Na⁺-binding residue glutamate-254 with an alanine residue. While the two crystal structures suggested an elevator-like movement to afford alternating access to the substrate binding site, the mechanistic role of Na⁺ and substrate in the conformational isomerization remains unclear. In this study, we utilized site-directed alkylation monitored by in-gel fluorescence (SDAF) to probe the solvent accessibility of the residues lining the substrate permeation pathway of ASBT_NM under different Na⁺ and substrate conditions, and interpreted the conformational states inferred from the crystal structures. Unexpectedly, the crosslinking experiments demonstrated that ASBT_NM is a monomer protein, unlike the other elevator-type transporters, usually forming a homodimer or a homotrimer. The conformational dynamics observed by the biochemical experiments were further validated using DEER measuring the distance between the spin-labelled pairs. Our results revealed that Na⁺ ions shift the conformational equilibrium of ASBT_NM toward the inward-facing state thereby facilitating cytoplasmic uptake of substrate. The current findings provide a novel perspective on the conformational equilibrium of secondary active transporters.     

The impacts of agricultural payments on groundwater quality: Spatial analysis on the case of Slovenia

The European Common Agricultural Policy still follows its primary goals, i.e. quality food at affordable prices and a decent standard of living for farmers, fifty years after its adoption. Moreover, this policy adapts to the changing needs of society and the new challenges, mostly preservation of the environment, nature and biodiversity in rural areas. Although the Common Agricultural Policy receives the largest share of European budget, the funds are decreasing over time, especially direct payments, which aim to provide basic income support to farmers in the European Union. On the other hand, agri-environmental payments are gaining importance. Policy decision-makers should be interested in the question of impacts of growing eco-conditionality of agricultural spending. New insights would help them to be successful in achieving the goals of sustainable agriculture. The purpose of this paper is to estimate the impacts of production support payments and rural development payments on the quality of groundwater. We use the small EU country Slovenia as an example. The baseline indicators are the level of nitrates and pesticides in groundwater, while the impacts were estimated using spatial error model. The results show that direct payments, coupled subsidies and investment grants raise the level of pesticides in groundwater, but do not have any statistically significant impact on the level of nitrates in groundwater. Furthermore, we did not find any statistically significant effects of agri-environmental payments on decrease of groundwater pollution with nitrates. However, our findings revealed that agri-environmental payments are effective in reducing pesticides in groundwater, although only to a limited extent. These results imply a problem of insufficient targeting of agri-environmental measures on the one hand, and suggest that greening of direct payments is necessary and entirely justified.     

Striking difference in response to expanding brood parasites by birds in western and eastern Beringia

Two species of obligate brood‐parasitic Cuculus cuckoos are expanding their ranges in Beringia. Both now breed on the Asian side, close to the Bering Strait, and are found in Alaska during the breeding season. From May to July 2017, we used painted 3D‐printed model eggs of two cuckoo host‐races breeding in northeastern Siberia to test behavioral responses of native songbirds on both sides of the Bering Strait, with particular attention to species that are known cuckoo hosts in their Siberian range. Each host nest was tested after the second egg was laid and, if possible, again 4 days later with a model of a different type. Although our Siberian study site was also outside the known breeding ranges of the cuckoos, we found that Siberian birds had strong anti‐parasite responses, with 14 of 22 models rejected. In contrast, birds in Alaska had virtually no detectable anti‐parasite behaviors, with only one of 96 models rejected; the rejecters were Red‐throated Pipits (Anthus cervinus). Such differences suggest that the cuckoos might successfully parasitize naïve hosts and become established in North America whether or not their historic host species are widely available.     

Total bile acids in hyperlipidaemic serum determined by bioluminescent and spectrophotometric methods

A simple, rapid and sensitive bioluminescent method has been used to measure total bile acids in hyperlipidaemic serum. We found that the levels of total bile acids in hypertriglyceridaemic and hypercholesterolemic sera determined by a spectrophotometric method were four-fold higher than those measured by the bioluminescent method (6.73 ± 4.07 μmol/l (mean ± SD) by bioluminescent and 26.10 ± 13.42 μmol/l by the spectrophotometric method). There was no difference in total bile acid levels between these two methods for normal serum (4.72 ± 3.38 μmol/l by bioluminescence and 4.49 ± 3.27 μmol/l by the spectrophotometric method).     

Investigations on the mechanism of hypercholesterolemia observed in copper deficiency in rats

The mechanism of hypercholesterolemia effect of Cu²⁺ deficiency was studied in rats. There was increased activity of hepatic hydroxymethylglutaryl-coenzyme A reductase and increased incorporation of labelled acetate into free cholesterol of liver in the Cu²⁺ deficient rats. Incorporation of label into ester cholesterol was however decreased in the liver. Concentration of bile acids in the liver was not significantly altered. Increase in the incorporation of labelled acetate into serum cholesterol and increase in the concentration of cholesterol and apo B in the low density lipoproteins + very low density lipoproteins fractions were observed. Activity of lipoprotein lipase of the extrahepatic tissues decreased in the Cu²⁺ deficient rats.     

我国某些常见化石硅藻的环境分析

本文详细研究了中国海表层沉积物中的常见化石硅藻,结果表明,这些种的分布范围及数量变化随环境的不同而变化。因此,根据柱状样品中这些种类数量的变化,能够推断古气候变化,恢复古地理环境,再现海平面波动历程,进而划分、对比第四系地层。     

普通小麦栽培品种丰抗13的4B—1D染色体易位的鉴定

通过细胞学观察,在普通小麦栽培品种“丰抗13”和“京红1号”的杂交后代中,发现有多价体出现,这就表明有染色体易位发生。为进一步弄清究竟是哪条染色体发生了易位,我们采用单体测交方法,观察鉴定所有各单体系F_1的花粉母细胞第一次减数分裂中期Ⅰ(以下简称PMCs中Ⅰ)染色体构型。从鉴定结果发现,凡2n=42的F_1 PMCs中Ⅰ出现19~Ⅱ 1~Ⅳ,而2n=41的F_1PMCs中Ⅰ的染色体构型不同,单体与易位有关的两个单体系4B和1D F_1 PMCs中的Ⅰ构型中有部分呈现为19个二价体加1个三价体,即19~Ⅱ 1~Ⅲ,没有单价体,而其余各单体系F_1 PMCs中Ⅰ构型则表现为18个二价体,1个四价体和1个单价体,即18~Ⅱ 1~Ⅰ 1~Ⅳ。因此,可以肯定“丰抗13”存在1个染色体易位,其有关染色体就是4B和1D。     

Immunological studies of ferredoxin-nitrite reductases and ferredoxin-glutamate synthases from photosynthetic organisms

Polyclonal antiserum specific for ferredoxin-nitrite reductase (EC 1.7.7.1) from the green alga Chlamydomonas reinhardii recognized the nitrite reductase from other green algae, but did not cross-react with the corresponding enzyme from different cyanobacteria or higher plant leaves. An analogous situation was also found for ferredoxin-glutamate synthase (EC 1.4.7.1), using its specific antiserum. Besides, the antibodies raised against C. reinhardii ferredoxin-glutamate synthase were able to inactivate the ferredoxin-dependent activity of nitrite reductase from green algae.These results suggest that there exist similar domains in ferredoxin-nitrite reductases and ferredoxin-glutamate synthases from green algae. In addition, both types of enzymes share common antigenic determinants, probably located at the ferredoxin-binding domain. In spite of their physicochemical resemblances, no apparent antigenic correlation exists between the corresponding enzymes from green algae and those from higher plant leaves or cyanobacteria.Abbreviations Fd ferredoxin - GOGAT glutamate synthase - MV⁺ reduced methyl viologen (radical cation) - NiR nitrite reductase - PMSF phenylmethylsulphonyl fluoride - SDS sodium dodecyl sulfate     

Ultrastructure of the kidney of a South American caecilian,Typhlonectes compressicaudus (Amphibia,Gymnophiona)

Summary The ultrastructure of the distal nephron, the collecting duct and the Wolffian duct was studied in a South American caecilian, Typhlonectes compressicaudus (Amphibia, Gymnophiona) by transmission and scanning electron microscopy (TEM, SEM). The distal tubule (DT) is made up of one type of cell that has a well-developed membrane labyrinth established both by interdigitating processes and by interlocking ramifications. The processes contain large mitochondria, the ramifications do not. The tight junction is shallow and elongated by a meandering course. The connecting tubule (CNT) is composed of CNT cells proper and intercalated cells, both of which are cuboidal in shape. The CNT cells are characterized by many lateral interlocking folds. The intercalated cells have a dark cytoplasm densely filled with mitochondria. Their apical cell membrane is typically amplified by microplicae beneath which a layer of globular particles (studs) is found. The collecting duct (CD) is composed of principal cells and intercalated cells, again both cuboidal in shape. The CD epithelium is characterized by dilated intercellular spaces, which are often filled with lateral microfolds projecting from adjacent principal cells. The apical membrane is covered by a prominent glycocalyx. The intercalated cells in the CD are similar to those in the CNT. The Wolffian duct (WD) has a tall pseudostratified epithelium established by WD cells proper, intercalated cells and basal cells. The WD cells contain irregular-shaped dense granules located beneath the apical cell membrane. The intercalated cells of the WD have a dark cytoplasm with many mitochondria; their nuclei display a dense chromatin pattern.Research fellow of the Alexander von Humboldt Foundation