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Amihai Karniel Devid Mrusek Wieland Steinchen Orly Dym Gert Bange Eitan Bibi 《Journal of molecular biology》2018,430(11):1607-1620
Much of our knowledge on the function of proteins is deduced from their mature, folded states. However, it is unknown whether partially synthesized nascent protein segments can execute biological functions during translation and whether their premature folding states matter. A recent observation that a nascent chain performs a distinct function, co-translational targeting in vivo, has been made with the Escherichia coli signal recognition particle receptor FtsY, a major player in the conserved pathway of membrane protein biogenesis. FtsY functions as a membrane-associated entity, but very little is known about the mode of its targeting to the membrane. Here we investigated the underlying structural mechanism of the co-translational FtsY targeting to the membrane. Our results show that helices N2–4, which mediate membrane targeting, form a stable folding intermediate co-translationally that greatly differs from its fold in the mature FtsY. These results thus resolve a long-standing mystery of how the receptor targets the membrane even when deleted of its alleged membrane targeting sequence. The structurally distinct targeting determinant of FtsY exists only co-translationally. Our studies will facilitate further efforts to seek cellular factors required for proper targeting and association of FtsY with the membrane. Moreover, the results offer a hallmark example for how co-translational nascent intermediates may dictate biological functions. 相似文献
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Nine monoclonal antibodies directed against class A beta-lactamases were detected and selected by a novel screening procedure based on assaying the modifications in the catalytic and stability properties of beta-lactamase in solution. Unlike conventional screening, e.g., ELISA or immunoprecipitation, the present method does not depend on firm binding and thus favors detection of low affinity antibodies. Individual antibodies were found to affect the enzymatic activity in various ways including stimulation, neutralization, protection and stabilization. Class A beta-lactamases show only 20% among members of this class. In contrast, two of our monoclonal antibodies cross-reacted with different beta-lactamases and thus demonstrate the presence of shared structural epitopes in this class of enzymes. One of the cross-reacting antibodies was elicited by sequential immunization with two different beta-lactamases. Taken together, our findings stress the importance of the screening method in antibody selection and illustrate the use of 'functional' monoclonal antibodies in the study of the structure-function relationship in an enzyme. 相似文献
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Parlitz R Eitan A Stjepanovic G Bahari L Bange G Bibi E Sinning I 《The Journal of biological chemistry》2007,282(44):32176-32184
Escherichia coli membrane protein biogenesis is mediated by a signal recognition particle and its membrane-associated receptor (FtsY). Although crucial for its function, it is still not clear how FtsY interacts with the membrane. Analysis of the structure/function differences between severely truncated active (NG+1) and inactive (NG) mutants of FtsY enabled us to identify an essential membrane-interacting determinant. Comparison of the three-dimensional structures of the mutants, combined with site-directed mutagenesis, modeling, and liposome-binding assays, revealed that FtsY contains a conserved autonomous lipid-binding amphipathic alpha-helix at the N-terminal end of the N domain. Deletion experiments showed that this helix is essential for FtsY function in vivo, thus offering, for the first time, clear evidence for the functionally important, physiologically relevant interaction of FtsY with lipids. 相似文献
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Shazia Iram Rabia Basri Khuram Shahzad Ahmad Shaan Bibi Jaffri 《Soil & Sediment Contamination》2019,28(4):411-430
Current investigation has for the first time utilized Trichocomaceae fungi i.e. Aspergillus niger, Aspergillus terreus, Aspergillus flavus and Pencillium i.e. Penicillium chrysogenum for augmenting the phytoremediation potential of bioenergy crops wheat (Zea mays) and ? sunflower (Helianthus annuus). Phytoremediation was done for mitigation of heavy metals i.e. Chromium (Cr), Copper (Cu), Lead (Pb) and Cadmium (Cd) from contaminated soils of agricultural significance. Phytoremediant crops were inoculated with fungal cultures by three methods i.e. mixing method, seed inoculation method and layering spreading method. Maize and sunflower plants after fungal inoculation were harvested after 60 days of germination. The estimation of % biomass and bioenergy of maize and sunflower plants was done. Results were indicative of the good phytoremediation potential of roots and shoots for uptake of heavy metals i.e. CrAspergillus niger, Aspergillus terreus and Aspergillus flavus by fungal inoculation methods. Sunflower and fungal inoculum of Aspergillus flavus and Penicillium chrysogenum extracted significant quantity of metals from the soil. By three fungal inoculation methods, range of % production of biomass was 84?87% and sunflower plants dry biomass 9.6 g yielded 0.16% of oil. Obtained results are have favored the use of fungal inoculation as an effective mode for phytoremediation augmentation of maize and sunflower. Furthermore, current work also signifies the sustainable conversion of bioenergy crops to biofuel production in a cost effective mode. 相似文献
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Evidence for coupling of membrane targeting and function of the signal recognition particle (SRP) receptor FtsY 总被引:3,自引:0,他引:3
Anat A. Herskovits Andrei Seluanov Ricardo Rajsbaum Corinne M. ten Hagen-Jongman Tanja Henrichs Elena S. Bochkareva Gregory J. Phillips Francis J. Probst Taiji Nakae Michael Ehrmann Joen Luirink Eitan Bibi 《EMBO reports》2001,2(11):1040-1046
Recent studies have indicated that FtsY, the signal recognition particle receptor of Escherichia coli, plays a central role in membrane protein biogenesis. For proper function, FtsY must be targeted to the membrane, but its membrane-targeting pathway is unknown. We investigated the relationship between targeting and function of FtsY in vivo, by separating its catalytic domain (NG) from its putative targeting domain (A) by three means: expression of split ftsY, insertion of various spacers between A and NG, and separation of A and NG by in vivo proteolysis. Proteolytic separation of A and NG does not abolish function, whereas separation by long linkers or expression of split ftsY is detrimental. We propose that proteolytic cleavage of FtsY occurs after completion of co-translational targeting and assembly of NG. In contrast, separation by other means may interrupt proper synchronization of co-translational targeting and membrane assembly of NG. The co-translational interaction of FtsY with the membrane was confirmed by in vitro experiments. 相似文献
60.
Elia Zomot Eliane Hadas Yardeni Attilio Vittorio Vargiu Heng-Keat Tam Giuliano Malloci Venkata Krishnan Ramaswamy Michal Perach Paolo Ruggerone Klaas Martinus Pos Eitan Bibi 《Journal of molecular biology》2018,430(9):1368-1385
Secondary multidrug (Mdr) transporters utilize ion concentration gradients to actively remove antibiotics and other toxic compounds from cells. The model Mdr transporter MdfA from Escherichia coli exchanges dissimilar drugs for protons. The transporter should open at the cytoplasmic side to enable access of drugs into the Mdr recognition pocket. Here we show that the cytoplasmic rim around the Mdr recognition pocket represents a previously overlooked important regulatory determinant in MdfA. We demonstrate that increasing the positive charge of the electrically asymmetric rim dramatically inhibits MdfA activity and sometimes even leads to influx of planar, positively charged compounds, resulting in drug sensitivity. Our results suggest that unlike the mutants with the electrically modified rim, the membrane-embedded wild-type MdfA exhibits a significant probability of an inward-closed conformation, which is further increased by drug binding. Since MdfA binds drugs from its inward-facing environment, these results are intriguing and raise the possibility that the transporter has a sensitive, drug-induced conformational switch, which favors an inward-closed state. 相似文献