共查询到20条相似文献,搜索用时 15 毫秒
1.
2.
3.
4.
5.
6.
The multidrug transporter, a double-edged sword 总被引:41,自引:0,他引:41
7.
The kinesin family member BimC has a highly positively charged domain of approximately 70 amino acids at the N terminus of the motor domain. Motor domain constructs of BimC were prepared with and without this extra domain to determine its influence. The level of microtubules needed for half saturation of the ATPase of BimC motor domain constructs is reduced by approximately 7000-fold at low ionic strength upon addition of this extra N-terminal extension. Although the change in microtubule affinity is less at higher salt, addition of the N-terminal domain still produces a 20-fold increase in affinity for microtubules in 200 mm potassium acetate. A fusion protein of the N-terminal domain and thioredoxin binds tightly to MTs at low salt, consistent with the increased affinity of motor domain constructs (which contain the N-terminal domain) being due to the additional binding of the N-terminal domain to the microtubule. Hydrodynamic analysis indicates that the N-terminal extension is in a highly extended conformation, suggesting that it may be intrinsically disordered. Fusion of the N-terminal extension of BimC onto the motor domain of conventional kinesin produces a similar large increase in microtubule affinity without significant reduction in kcat or velocity in an in vitro motility assay, suggesting that the N-terminal extension can act in a modular manner to increase the microtubule affinity of kinesin motor domains without a decrease in velocity. 相似文献
8.
9.
Cloning of the human thiamine transporter, a member of the folate transporter family. 总被引:7,自引:0,他引:7
B Dutta W Huang M Molero R Kekuda F H Leibach L D Devoe V Ganapathy P D Prasad 《The Journal of biological chemistry》1999,274(45):31925-31929
We have isolated a cDNA from human placenta, which, when expressed heterologously in mammalian cells, mediates the transport of the water-soluble vitamin thiamine. The cDNA codes for a protein of 497 amino acids containing 12 putative transmembrane domains. Northern blot analysis indicates that this transporter is widely expressed in human tissues. When expressed in HeLa cells, the cDNA induces the transport of thiamine (K(t) = 2.5 +/- 0.6 microM) in a Na(+)-independent manner. The cDNA-mediated transport of thiamine is stimulated by an outwardly directed H(+) gradient. Substrate specificity assays indicate that the transporter is specific to thiamine. Even though thiamine is an organic cation, the cDNA-induced thiamine transport is not inhibited by other organic cations. Similarly, thiamine is not a substrate for the known members of mammalian organic cation transporter family. The thiamine transporter gene, located on human chromosome 1q24, consists of 6 exons and is most likely the gene defective in the metabolic disorder, thiamine-responsive megaloblastic anemia. At the level of amino acid sequence, the thiamine transporter is most closely related to the reduced-folate transporter and thus represents the second member of the folate transporter family. 相似文献
10.
The 2-hydroxycarboxylate transporter family is a family of secondary transporters found exclusively in the bacterial kingdom. They function in the metabolism of the di- and tricarboxylates malate and citrate, mostly in fermentative pathways involving decarboxylation of malate or oxaloacetate. These pathways are found in the class Bacillales of the low-CG gram-positive bacteria and in the gamma subdivision of the Proteobacteria. The pathways have evolved into a remarkable diversity in terms of the combinations of enzymes and transporters that built the pathways and of energy conservation mechanisms. The transporter family includes H+ and Na+ symporters and precursor/product exchangers. The proteins consist of a bundle of 11 transmembrane helices formed from two homologous domains containing five transmembrane segments each, plus one additional segment at the N terminus. The two domains have opposite orientations in the membrane and contain a pore-loop or reentrant loop structure between the fourth and fifth transmembrane segments. The two pore-loops enter the membrane from opposite sides and are believed to be part of the translocation site. The binding site is located asymmetrically in the membrane, close to the interface of membrane and cytoplasm. The binding site in the translocation pore is believed to be alternatively exposed to the internal and external media. The proposed structure of the 2HCT transporters is different from any known structure of a membrane protein and represents a new structural class of secondary transporters. 相似文献
11.
Evidence for auto-inhibition by the N terminus of hADAR2 and activation by dsRNA binding 总被引:1,自引:0,他引:1
Adenosine deaminases that act on RNA (ADARs) catalyze adenosine to inosine conversion in RNA that is largely double stranded. Human ADAR2 (hADAR2) contains two double-stranded RNA binding motifs (dsRBMs), separated by a 90-amino acid linker, and these are followed by the C-terminal catalytic domain. We assayed enzymatic activity of N-terminal deletion constructs of hADAR2 to determine the role of the dsRBMs and the intervening linker peptide. We found that a truncated protein consisting of one dsRBM and the deaminase domain was capable of deaminating a short 15-bp substrate. In contrast, full-length hADAR2 was inactive on this short substrate. In addition, we observed that the N terminus, which was deleted from the truncated protein, inhibits editing activity when added in trans. We propose that the N-terminal domain of hADAR2 contains sequences that cause auto-inhibition of the enzyme. Our results suggest activation requires binding to an RNA substrate long enough to accommodate interactions with both dsRBMs. 相似文献
12.
Ivan Birukou Susan M. Seo Bryan D. Schindler Glenn W. Kaatz Richard G. Brennan 《Nucleic acids research》2014,42(4):2774-2788
The multidrug efflux pump MepA is a major contributor to multidrug resistance in Staphylococcus aureus. MepR, a member of the multiple antibiotic resistance regulator (MarR) family, represses mepA and its own gene. Here, we report the structure of a MepR–mepR operator complex. Structural comparison of DNA-bound MepR with ‘induced’ apoMepR reveals the large conformational changes needed to allow the DNA-binding winged helix-turn-helix motifs to interact with the consecutive major and minor grooves of the GTTAG signature sequence. Intriguingly, MepR makes no hydrogen bonds to major groove nucleobases. Rather, recognition-helix residues Thr60, Gly61, Pro62 and Thr63 make sequence-specifying van der Waals contacts with the TTAG bases. Removing these contacts dramatically affects MepR–DNA binding activity. The wings insert into the flanking minor grooves, whereby residue Arg87, buttressed by Asp85, interacts with the O2 of T4 and O4′ ribosyl oxygens of A23 and T4. Mutating Asp85 and Arg87, both conserved throughout the MarR family, markedly affects MepR repressor activity. The His14′:Arg59 and Arg10′:His35:Phe108 interaction networks stabilize the DNA-binding conformation of MepR thereby contributing significantly to its high affinity binding. A structure-guided model of the MepR–mepA operator complex suggests that MepR dimers do not interact directly and cooperative binding is likely achieved by DNA-mediated allosteric effects. 相似文献
13.
14.
Defining the mechanism of activation of AMP-activated protein kinase by the small molecule A-769662, a member of the thienopyridone family 总被引:1,自引:0,他引:1
Sanders MJ Ali ZS Hegarty BD Heath R Snowden MA Carling D 《The Journal of biological chemistry》2007,282(45):32539-32548
AMP-activated protein kinase (AMPK) plays a key role in maintaining energy homeostasis. Activation of AMPK in peripheral tissues has been shown to alleviate the symptoms of metabolic diseases, such as type 2 diabetes, and consequently AMPK is a target for treatment of these diseases. Recently, a small molecule activator (A-769662) of AMPK was identified that had beneficial effects on metabolism in ob/ob mice. Here we show that A-769662 activates AMPK both allosterically and by inhibiting dephosphorylation of AMPK on Thr-172, similar to the effects of AMP. A-769662 activates AMPK harboring a mutation in the gamma subunit that abolishes activation by AMP. An AMPK complex lacking the glycogen binding domain of the beta subunit abolishes the allosteric effect of A-769662 but not the allosteric activation by AMP. Moreover, mutation of serine 108 to alanine, an autophosphorylation site within the glycogen binding domain of the beta1 subunit, almost completely abolishes activation of AMPK by A-769662 in cells and in vitro, while only partially reducing activation by AMP. Based on our results we propose a model for activation of AMPK by A-769662. Importantly, this model may provide clues for understanding the mechanism by which AMP leads to activation of AMPK, which in turn may help in the identification of other AMPK activators. 相似文献
15.
16.
AtOPT3, a member of the oligopeptide transporter family,is essential for embryo development in Arabidopsis 总被引:1,自引:0,他引:1
下载免费PDF全文

A T-DNA-tagged population of Arabidopsis was screened for mutations in AtOPT3, which encodes a member of the oligopeptide (OPT) family of peptide transporters, and a recessive mutant allele, opt3, was identified. Phenotypic analysis of opt3 showed that most homozygous embryos were arrested at or before the octant stage of embryo development and that none showed the usual periclinal division leading to the formation of the protoderm. This defective phenotype could be reversed by complementation with the full-length, wild-type AtOPT3 gene. A beta-glucuronidase (GUS) fusion to DNA sequences upstream of the putative AtOPT3 ATG start codon was constructed, and the expression pattern was assayed in transgenic plants. AtOPT3 was expressed in the vascular tissues of seedlings and mature plants as well as in pollen. Consistent with the function of AtOPT3 in embryogenesis, AtOPT3::GUS expression also was detected in developing embryos and in the maternal tissues of seeds. These data suggest a critical role for peptide transport in early embryo development. 相似文献
17.
18.
19.
Multidrug resistance is a serious problem in successful cancer chemotherapy. Studies using model cell lines have demonstrated that overexpression of some members of the ATP-binding cassette (ABC) transporter superfamily, such as ABCC1, causes enhanced efflux and, thus, decreased accumulation of multiple anticancer drugs, which leads to increased cell survival. Unlike most other ABC transporters, ABCC1 has an additional membrane-spanning domain (MSD0) with a putative extracellular amino terminus of 32 amino acids. However, the function of MSD0 and the role of the extracellular amino terminus are largely unknown. In this study, we examined the structural folding and the function of the amino terminus. We found that it has a U-shaped folding with the bottom of the U-structure facing cytoplasm and both ends in extracellular space. We also found that this U-shaped amino terminus probably functions as a gate to regulate the drug transport activity of human ABCC1. 相似文献