The C Terminus of the Alb3 Membrane Insertase Recruits cpSRP43 to the Thylakoid Membrane

The YidC/Oxa1/Alb3 family of membrane proteins controls the insertion and assembly of membrane proteins in bacteria, mitochondria, and chloroplasts. Here we describe the molecular mechanisms underlying the interaction of Alb3 with the chloroplast signal recognition particle (cpSRP). The Alb3 C-terminal domain (A3CT) is intrinsically disordered and recruits cpSRP to the thylakoid membrane by a coupled binding and folding mechanism. Two conserved, positively charged motifs reminiscent of chromodomain interaction motifs in histone tails are identified in A3CT that are essential for the Alb3-cpSRP43 interaction. They are absent in the C-terminal domain of Alb4, which therefore does not interact with cpSRP43. Chromodomain 2 in cpSRP43 appears as a central binding platform that can interact simultaneously with A3CT and cpSRP54. The observed negative cooperativity of the two binding events provides the first insights into cargo release at the thylakoid membrane. Taken together, our data show how Alb3 participates in cpSRP-dependent membrane targeting, and our data provide a molecular explanation why Alb4 cannot compensate for the loss of Alb3. Oxa1 and YidC utilize their positively charged, C-terminal domains for ribosome interaction in co-translational targeting. Alb3 is adapted for the chloroplast-specific Alb3-cpSRP43 interaction in post-translational targeting by extending the spectrum of chromodomain interactions.     

Investigation of the cumulative tissue doses of naphthoquinones in human serum using protein adducts as biomarker of exposure

Both 1,2-naphthoquinone (1,2-NPQ) and 1,4-naphthoquinone (1,4-NPQ) are reactive metabolites of naphthalene that are thought to be responsible for the naphthalene-induced cytotoxicity and genotoxicity. The aim of this study was to investigate the cumulative tissue dose of 1,2-NPQ and 1,4-NPQ in human serum derived from blood donors in Taiwan via measurements of albumin adducts by a methodology, which employs trifluoroacetic acid anhydride and methanesulfonic acid to selectively cleave cysteinyl adducts on proteins. Both 1,2-NPQ and 1,4-NPQ adducts were detected in all male and female subjects (n = 22). The median levels of 1,2-NPQ adduct in human subjects were estimated to be 268 (range 139-857) and 203 (range 128-1352) (pmol/g) in male (n = 11) and female (n = 11) subjects, respectively. In contrast, the median levels of 1,4-NPQ adduct were estimated to be 45.0 (range 22.0-117) and 38.9 (range 21.5-172) (pmol/g) in male and female subjects, respectively. We noticed that levels of 1,2-NPQ adduct were significantly correlated with those of 1,4-NPQ adduct (correlation coefficient r = 0.643, p < 0.01). Results from in vitro experiments confirmed that the production of naphthoquinones-derived adducts on serum albumin increased with increased concentration of naphthoquinones (0-100 μM). Linear relationships were observed over the range of concentration. Time-course experiments suggested that both 1,2-NPQ and 1,4-NPQ-derived adducts rapidly reached maximum values at 10 min mark and remained constant thereafter. The reaction rate constant analyses indicated that the second-order rate constants, representing in vitro reactions between naphthoquinones and cysteine residues of serum albumin, were estimated to be 0.0044/0.0002 L(g protein)⁻¹ h⁻¹, respectively. Overall, the cumulative tissue doses of 1,4-NPQ (217-316 nM h) in male and female subjects were ∼3-fold greater than those of 1,2-NPQ (76-98 nM h) in the study population. The initial concentrations of serum 1,2-NPQ and 1,4-NPQ in the study population were estimated to be between 145-188 and 807-1175 nM, respectively. We conclude that the relatively large amounts of naphthoquinones present in human serum may point to toxicological consequences.     

2-Mercapto-5-benzimidazolesulfonic acid: an effective multimodal ligand for the separation of antibodies

The report describes the use of 2-mercapto-5-benzimidazolesulfonic acid (MBISA) as a ligand for the separation of antibodies by chromatography. The ligand shows a relatively specific adsorption property for antibodies from very crude biologicals at pH 5.0-5.5. At this pH range most of other proteins do not interact with the resin especially when the ionic strength is similar to physiological conditions. Several characterization studies are described such as antibody adsorption in different conditions of ionic strength, pH and temperature. These properties are advantageously used to selectively capture antibodies from very crude feed stocks without dilution or addition of lyotropic salts. Demonstration was made that the adsorption mechanism is neither based on ion exchange nor on hydrophobic associations, but rather as an assembly of a variety of properties of the ligand itself. Binding capacity in the described conditions ranges between 25 and 30 mg/mL of resin. The sorbent does not co-adsorb albumin (Alb) and seems compatible with a large variety of feedstocks. Quantitative antibody desorption occurs when the pH is raised above 8.5. The final purity of the antibody depends on the nature of the feedstock, and can reach levels of purity as high as 98%. Even with very crude biological liquids such as ascites fluids, cell culture supernatants and Chon fraction II + III from human plasma fractionation where the number of protein impurities is particularly large, immunoglobumins G (IgG) were separated at high purity level in a single step.     

振幅整合脑电图评分联合ox-LDL/HDL比值、hs-CRP/Alb比值、FAR对急性脑梗死患者预后的预测价值

摘要 目的：探讨振幅整合脑电图（aEEG）评分联合氧化低密度脂蛋白/高密度脂蛋白（ox-LDL/HDL）比值、高敏C反应蛋白/白蛋白（hs-CRP/Alb）比值、纤维蛋白原/白蛋白比值（FAR）对急性脑梗死（ACI）患者预后的预测价值。方法：选择2021年1月至2022年l2月我院收治的242例ACI患者,均行aEEG检查获得aEEG评分,检测血清ox-LDL/HDL比值、hs-CRP/Alb比值、FAR。出院3个月后采用改良Rankin量表（mRS）评估患者预后。收集ACI患者的临床资料,多因素Logistic回归分析ACI患者预后不良的影响因素。受试者工作用特征（ROC）曲线分析aEEG评分、血清ox-LDL/HDL比值、hs-CRP/Alb比值、FAR预测ACI患者预后的效能。结果：预后不良组aEEG评分、血清ox-LDL/HDL比值、hs-CRP/Alb比值、FAR均高于预后良好组（P＜0.05）。多发脑梗死病灶、高NHISS评分、高aEEG评分、高ox-LDL/HDL比值、高hs-CRP/Alb比值和高FAR是ACI患者预后不良的危险因素（P＜0.05）。aEEG评分、血清ox-LDL/HDL比值、hs-CRP/Alb比值、FAR预测ACI患者预后的曲线下面积分别为0.793、0.759、0.777、0.821,联合预测的曲线下面积为0.915,高于单独指标预测。结论：ACI预后不良患者aEEG评分、ox-LDL/HDL比值、hs-CRP/Alb比值、FAR均升高,上述指标联合应用预测ACI患者预后不良具有较高的价值。     

The Role of the Strictly Conserved Positively Charged Residue Differs among the Gram-positive,Gram-negative,and Chloroplast YidC Homologs

Recently, the structure of YidC2 from Bacillus halodurans revealed that the conserved positively charged residue within transmembrane segment one (at position 72) is located in a hydrophilic groove that is embedded in the inner leaflet of the lipid bilayer. The arginine residue was essential for the Bacillus subtilis SpoIIIJ (YidC1) to insert MifM and to complement a SpoIIIJ mutant strain. Here, we investigated the importance of the conserved positively charged residue for the function of the Escherichia coli YidC, Streptococcus mutans YidC2, and the chloroplast Arabidopsis thaliana Alb3. Like the Gram-positive B. subtilis SpoIIIJ, the conserved arginine was required for functioning of the Gram-positive S. mutans YidC2 and was necessary to complement the E. coli YidC depletion strain and to promote insertion of a YidC-dependent membrane protein synthesized with one but not two hydrophobic segments. In contrast, the conserved positively charged residue was not required for the E. coli YidC or the A. thaliana Alb3 to functionally complement the E. coli YidC depletion strain or to promote insertion of YidC-dependent membrane proteins. Our results also show that the C-terminal half of the helical hairpin structure in cytoplasmic loop C1 is important for the activity of YidC because various deletions in the region either eliminate or impair YidC function. The results here underscore the importance of the cytoplasmic hairpin region for YidC and show that the arginine is critical for the tested Gram-positive YidC homolog but is not essential for the tested Gram-negative and chloroplast YidC homologs.     

慢性肾小球肾炎患者治疗前后血清CRP和VEGF浓度变化及临床意义

目的:研究慢性肾小球肾炎(CNG)中血清C反应蛋白(CRP)和血管内皮生长因子(VEGF)的浓度变化及其临床意义。方法:采用ELISA法测定35例正常对照组与41例慢性肾小球肾炎患者治疗前后血清IL-6和VEGF的浓度,同时放射免疫分析法测定血清TNF-α浓度,免疫比浊法测定血清CRP与尿Alb浓度。结果:①治疗前后CNG患者血清中IL-6、TNF-α和CRP较正常对照组均显著升高(P<0.05或P<0.01),但治疗后IL-6、TNF-α和CRP水平显著低于治疗前(P<0.01),且血清CRP与IL-6和TNF-α呈正相关(P<0.01)。②治疗后,CNG患者血清VEGF水平与尿Alb含量较治疗前明显降低(P<0.05或P<0.01),仍显著高于正常对照组(P<0.05或P<0.01),且血清VEGF与尿Alb水平呈正相关(P<0.01)。结论:CRP、IL-6和TNF-α参与了CNG患者慢性炎症反应,VEGF则与蛋白尿的产生密切相关,治疗前后血清CRP和VEGF检测对于慢性肾小球肾炎的病情了解及临床疗效评估均具有重要的临床价值。     

不同危险分层急性肺栓塞患者D-二聚体与纤维蛋白原比值、中性粒细胞与淋巴细胞比值、白蛋白的变化及其与预后的关系研究

摘要 目的：探讨不同危险分层急性肺栓塞（APE）患者D-二聚体与纤维蛋白原比值（DFR）、中性粒细胞与淋巴细胞比值（NLR）、白蛋白（Alb）的变化及其与预后的关系。方法：选择2019年3月至2021年12月我院收治的APE患者154例作为APE组,根据《肺血栓栓塞症的诊断与治疗指南(2015)》分为低危组48例、中危组69例和高危组37例,另选择同期我院体检健康志愿者40例作为对照组,比较各组DFR、NLR、Alb水平。根据不同预后将APE患者分为存活组125例,死亡组29例,比较两组DFR、NLR、Alb水平。应用受试者工作特征（ROC）曲线分析DFR、NLR、Alb对APE预后的预测价值。结果：APE组DFR、NLR显著高于对照组,Alb水平显著低于对照组（P<0.05）。随着危险分层增加,APE患者DFR、NLR逐渐升高,Alb水平逐渐降低,不同危险分层APE患者DFR、NLR、Alb水平比较有统计学意义（P<0.05）。死亡组DFR、NLR显著高于存活组,Alb水平显著低于存活组（P<0.05）。ROC曲线分析显示,DFR、NLR、Alb对APE死亡预测具有较高的敏感度、特异度,其中DFR、NLR、Alb联合检测对APE死亡预测的曲线下面积（AUC）、敏感度、特异度最高。结论：APE患者DFR、NLR异常升高,Alb异常降低与APE危险分层增加及不良预后相关,DFR、NLR、Alb联合检测对APE患者预后不良的预测价值更高。     

Brugia malayi: Effects of nitazoxanide and tizoxanide on adult worms and microfilariae of filarial nematodes

There is an urgent need for safe and effective antifilarials. Prior studies have shown that the nitazoxanide (NTZ) exhibits broad activity against anaerobic bacteria, protozoa, and certain intestinal helminths. We examined the effects of NTZ and tizoxanide (TZ) on Brugia malayi nematodes in vitro and in vivo. In vitro, NTZ and TZ reduced worm motility and viability in a dose-dependent manner. Worm viability was reduced by 50% with both compounds at 2.5 and 20 μg/ml killed adult worms. NTZ or TZ (5 μg/ml) significantly reduced microfilaria release. These compounds blocked worm’s embryogenesis, and decreased microfilarial motility and viability. Treated worms had damaged cuticles and abnormal mitochondria. Wolbachia were not cleared by NTZ or TZ treatment. Neither NTZ nor TZ cleared adult worms or microfilariae in infected gerbils. These results show that NTZ and TZ have potent effects on B. malayi nematodes in vitro. However, they were not effective in vivo.