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1.
The integral membrane proteins of the ammonium transporter (AMT/Rh) family provide the major route for shuttling ammonium (NH(4)(+)/NH(3)) across bacterial, archaeal, fungal and plant membranes. These proteins are distantly related to the Rh (rhesus) glycoproteins, which are absent in higher plants, but are present in many species, including bacteria and mammals. It appears that the large nitrogen requirement of plants resulted in unique strategies to acquire, capture and/or release ammonium. The biological function of plant ammonium transporters will be discussed and compared to other AMT/Rh proteins. 相似文献
2.
Monahan BJ Unkles SE Tsing I T Kinghorn JR Hynes MJ Davis MA 《Fungal genetics and biology : FG & B》2002,36(1):35-46
The movement of ammonium across biological membranes is mediated in both prokaryotic and eukaryotic systems by ammonium transport proteins which constitute a family of related sequences (called the AMT/MEP family). Interestingly, recent evidence suggests that human and mouse Rhesus proteins which display significant relatedness to AMT/MEP sequences may function as ammonium transporters. To add to the functional understanding of ammonium transport proteins, the sequence changes in 37 loss-of-function mutations within the Aspergillus nidulans ammonium permease gene, meaA, were characterized. Together with the identification of conserved AMT/MEP residues and regions, the mutational analysis predicted regions important for uptake activity. Specifically, a major facilitator superfamily like motif (161-GAVAERGR-168 in MeaA) may be important for the translocation of ammonium across the membrane as may the conserved Pro186 residue. A specific Gly447 to Asp mutation was introduced into MeaA and this mutant protein was found to trans-inhibit the activity of endogenous MeaA and the other A. nidulans ammonium transporter, MepA. These results suggest that MeaA may interact with itself and with MepA, although any hetero-interaction is not required for ammonium transport function. In addition, cross-feeding studies showed that MeaA and to a lesser extent MepA are also required for the retention of intracellular ammonium. 相似文献
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Functional and physiological evidence for a rhesus-type ammonia transporter in Nitrosomonas europaea
Weidinger K Neuhäuser B Gilch S Ludewig U Meyer O Schmidt I 《FEMS microbiology letters》2007,277(2):260-267
Ammonium transporters form a conserved family of transport proteins and are widely distributed among all domains of life. The genome of Nitrosomonas europaea codes for a single gene (rh1) that belongs to the family of the AMT/Rh ammonium transporters. For the first time, this study provides functional and physiological evidence for a rhesus-type ammonia transporter in bacteria (N. europaea). The methylammonium (MA) transport activity of N. europaea correlated with the Rh1 expression. The K(m) value for the MA uptake of N. europaea was 1.8+/-0.2 mM (pH 7.25), and the uptake was competitively inhibited by ammonium [K(i)(NH(4) (+)) 0.3+/-0.1 mM at pH 7.25]. The MA uptake rate was pH dependent, indicating that the uncharged form of MA is transported by Rh1. An effect of the glutamine synthetase on the MA uptake was not observed. When expressed in Saccharomyces cerevisiae, the function of Rh1 from N. europaea as an ammonia/MA transporter was confirmed. The results suggest that Rh1 equilibrates the uncharged substrate species. A low pH value in the periplasmic space during ammonia oxidation seems to be responsible for the ammonium accumulation functioning as an acid NH(4) (+) trap. 相似文献
5.
The transport of ammonium/ammonia is a key process for the acquisition and metabolism of nitrogen. Ammonium transport is mediated by the AMT/MEP/Rh family of membrane proteins which are found in microorganisms, plants, and animals, including the Rhesus blood group antigens in humans. Although ammonium transporters from all kingdoms have been functionally expressed and partially characterized, the transport mechanism, as well as the identity of the true substrate (NH(4+) or NH(3)) remains unclear. Here we describe the functional expression and characterization of LeAMT1;1, a root hair ammonium transporter from tomato (Lycopersicon esculentum) in Xenopus oocytes. Micromolar concentrations of external ammonium were found to induce concentration- and voltage-dependent inward currents in oocytes injected with LeAMT1;1 cRNA, but not in water-injected control oocytes. The NH(4+)-induced currents were more than 3-fold larger than methylammonium currents and were not subject to inhibition by Na(+) or K(+). The voltage dependence of the affinity of LeAMT1;1 toward its substrate strongly suggests that charged NH(4+), rather than NH(3), is the true transport substrate. Furthermore, ammonium transport was independent of the external proton concentration between pH 5.5 and pH 8.5. LeAMT1;1 is concluded to mediate potential-driven NH(4+) uptake and retrieval depending on root membrane potential and NH(4+) concentration gradient. 相似文献
6.
Ortiz-Ramirez C Mora SI Trejo J Pantoja O 《The Journal of biological chemistry》2011,286(36):31113-31122
One of the main forms of nitrogen assimilated by microorganisms and plants is ammonium, despite its toxicity at low millimolar concentrations. Ammonium absorption has been demonstrated to be carried out by highly selective plasma membrane-located transporters of the AMT/MEP/Rh family and characterized by the presence of a well conserved hydrophobic pore through which ammonia is proposed to move. However, uncertainties exist regarding the exact chemical species transported by these membrane proteins, which can be in the form of either hydrophobic ammonia or charged ammonium. Here, we present the characterization of PvAMT1;1 from the common bean and demonstrate that it mediates the high affinity (micromolar), rapidly saturating (1 mM) electrogenic transport of ammonium. Activity of the transporter is enhanced by low extracellular pH, and associated with this acidic pH stimulation are changes in the reversal potential and cytoplasm acidification, indicating that PvAMT1;1 functions as an H(+)/NH(4)(+) symporter. Mutation analysis of a unique histidine present in PvAMT1;1 (H125R) leads to the stimulation of ammonium transport by decreasing the K(m) value by half and by increasing the V(max) 3-fold, without affecting the pH dependence of the symporter. In contrast, mutation of the first conserved histidine within the channel modifies the properties of PvAMT1;1, increasing its K(m) and V(max) values and transforming it into a pH-independent mechanism. 相似文献
7.
The molecular physiology of ammonium uptake and retrieval 总被引:18,自引:0,他引:18
Plants are able to take up ammonium from the soil, or through symbiotic interactions with microorganisms, via the root system. Using functional complementation of yeast mutants, it has been possible to identify a new class of membrane proteins, the ammonium transporter/methylammonium permease (AMT/MEP) family, that mediate secondary active ammonium uptake in eukaryotic and prokaryotic organisms. In plants, the AMT gene family can be subdivided according to their amino-acid sequences into three subfamilies: a large subfamily of AMT1 genes and two additional subfamilies each with single members (LeAMT1;3 from tomato and AtAMT2;1 from Arabidopsis thaliana). These transporters vary especially in their kinetic properties and regulatory mechanism. High-affinity transporters are induced in nitrogen-starved roots, whereas other transporters may be considered as the 'work horses' that are active when conditions are conducive to ammonium assimilation. The expression of several AMTs in root hairs further supports a role in nutrient acquisition. These studies provide basic information that will be needed for the dissection of nitrogen uptake by plants at the molecular level and for determining the role of individual AMTs in nutrient uptake and potentially in nutrient efficiency. 相似文献
8.
Investigating the phylogenetic relationships within physiologically essential gene families across a broad range of taxa can reveal the key gene duplication events underlying their family expansion and is thus important to functional genomics studies. P-Type II ATPases represent a large family of ATP powered transporters that move ions across cellular membranes and includes Na+/K+ transporters, H+/K+ transporters, and plasma membrane Ca2+ pumps. Here, we examine the evolutionary history of one such transporter, the Sarco(endo)plasmic reticulum calcium ATPase (SERCA), which maintains calcium homeostasis in the cell by actively pumping Ca2+ into the sarco(endo)plasmic reticulum. Our protein-based phylogenetic analyses across Eukaryotes revealed two monophyletic clades of SERCA proteins, one containing animals, fungi, and plants, and the other consisting of plants and protists. Our analyses suggest that the three known SERCA proteins in vertebrates arose through two major gene duplication events after the divergence from tunicates, but before the separation of fishes and tetrapods. In plants, we recovered two SERCA clades, one being the sister group to Metazoa and the other to Apicomplexa clade, suggesting an ancient duplication in an early eukaryotic ancestor, followed by subsequent loss of one copy in Opisthokonta, the other in protists, and retention of both in plants. We also report relatively recent and independent gene duplication events within invertebrate taxa including tunicates and the leech Helobdella robusta. Thus, it appears that both ancient and recent gene duplication events have played an important role in the evolution of this ubiquitous gene family across the eukaryotic domain. 相似文献
9.
Although theoretical studies have suggested that base-compositional heterogeneity can adversely affect phylogenetic reconstruction, only a few empirical examples of this phenomenon, mostly among ancient lineages (with divergence dates > 100 Mya), have been reported. In the course of our phylogenetic research on the New World marsupial family Didelphidae, we sequenced 2790 bp of the RAG1 exon from exemplar species of most extant genera. Phylogenetic analysis of these sequences recovered an anomalous node consisting of two clades previously shown to be distantly related based on analyses of other molecular data. These two clades show significantly increased GC content at RAG1 third codon positions, and the resulting convergence in base composition is strong enough to overwhelm phylogenetic signal from other genes (and morphology) in most analyses of concatenated datasets. This base-compositional convergence occurred relatively recently (over tens rather than hundreds of millions of years), and the affected gene region is still in a state of evolutionary disequilibrium. Both mutation rate and substitution rate are higher in GC-rich didelphid taxa, observations consistent with RAG1 sequences having experienced a higher rate of recombination in the convergent lineages. 相似文献
10.
《Channels (Austin, Tex.)》2013,7(6):496-502
In organisms from all kingdoms of life, ammonia and its conjugated ion ammonium are transported across membranes by proteins of the AMT/Rh family. Efficient and successful growth often depends on sufficient ammonium nutrition. The proteins mediating this transport, the so called Ammonium Transporter (AMT) or Rhesus like (Rh) proteins, share a very similar trimeric overall structure and a high sequence similarity even throughout the kingdoms. Even though structural components of the transport mechanism, like an external substrate recruitment site, an essential twin histidine pore motif, a phenylalanine gate and the hydrophobic pore are strongly conserved and have been analyzed in detail by molecular dynamic simulations and mutational studies, the substrate(s), which pass the central pores of the AMT/Rh subunits, NH4+, NH3 + H+, NH4+ + H+ or NH3, are still a matter of debate for most proteins, including the best characterized AmtB protein from Escherichia coli. The lack of a robust expression system for functional analysis has hampered proof of structural and mutational studies, although the NH3 transport function for Rh-like proteins is rarely disputed. In plant transporters belonging to the subfamily AMT1, transport is associated with electrical currents, while some plant transporters, notably of the AMT2 type, were suggested to transport NH3 across the membrane, without associated ionic currents. Here we summarize data in favor of each substrate for the distinct AMT/Rh classes, discuss mutants and how they differ in structure and functionality. A common mechanism with deprotonation and subsequent NH3 transport through the central subunit pore is suggested. 相似文献
11.
In organisms from all kingdoms of life, ammonia and its conjugated ion ammonium are transported across membranes by proteins of the AMT/Rh family. Efficient and successful growth often depends on sufficient ammonium nutrition. The proteins mediating this transport, the so called Ammonium Transporter (AMT) or Rhesus like (Rh) proteins, share a very similar trimeric overall structure and a high sequence similarity even throughout the kingdoms. Even though structural components of the transport mechanism, like an external substrate recruitment site, an essential twin histidine pore motif, a phenylalanine gate and the hydrophobic pore are strongly conserved and have been analyzed in detail by molecular dynamic simulations and mutational studies, the substrate(s), which pass the central pores of the AMT/Rh subunits, NH4+, NH3 + H+, NH4+ + H+ or NH3, are still a matter of debate for most proteins, including the best characterized AmtB protein from Escherichia coli. The lack of a robust expression system for functional analysis has hampered proof of structural and mutational studies, although the NH3 transport function for Rh-like proteins is rarely disputed. In plant transporters belonging to the subfamily AMT1, transport is associated with electrical currents, while some plant transporters, notably of the AMT2 type, were suggested to transport NH3 across the membrane, without associated ionic currents. Here we summarize data in favor of each substrate for the distinct AMT/Rh classes, discuss mutants and how they differ in structure and functionality. A common mechanism with deprotonation and subsequent NH3 transport through the central subunit pore is suggested. 相似文献
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Ludewig U Wilken S Wu B Jost W Obrdlik P El Bakkoury M Marini AM André B Hamacher T Boles E von Wirén N Frommer WB 《The Journal of biological chemistry》2003,278(46):45603-45610
In most organisms, high affinity ammonium uptake is catalyzed by members of the ammonium transporter family (AMT/MEP/Rh). A single point mutation (G458D) in the cytosolic C terminus of the plasma membrane transporter LeAMT1;1 from tomato leads to loss of function, although mutant and wild type proteins show similar localization when expressed in yeast or plant protoplasts. Co-expression of LeAMT1;1 and mutant in Xenopus oocytes inhibited ammonium transport in a dominant negative manner, suggesting homo-oligomerization. In vivo interaction between LeAMT1;1 proteins was confirmed by the split ubiquitin yeast two-hybrid system. LeAMT1;1 is isolated from root membranes as a high molecular mass oligomer, converted to a approximately 35-kDa polypeptide by denaturation. To investigate interactions with the LeAMT1;2 paralog, co-localizing with LeAMT1;1 in root hairs, LeAMT1;2 was characterized as a lower affinity NH4+ uniporter. Co-expression of wild types with the respective G458D/G465D mutants inhibited ammonium transport in a dominant negative manner, supporting the formation of heteromeric complexes in oocytes. Thus, in yeast, oocytes, and plants, ammonium transporters are able to oligomerize, which may be relevant for regulation of ammonium uptake. 相似文献
14.
Evolution and functional characterization of the RH50 gene from the ammonia-oxidizing bacterium Nitrosomonas europaea 总被引:1,自引:0,他引:1
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Cherif-Zahar B Durand A Schmidt I Hamdaoui N Matic I Merrick M Matassi G 《Journal of bacteriology》2007,189(24):9090-9100
The family of ammonia and ammonium channel proteins comprises the Amt proteins, which are present in all three domains of life with the notable exception of vertebrates, and the homologous Rh proteins (Rh50 and Rh30) that have been described thus far only in eukaryotes. The existence of an RH50 gene in bacteria was first revealed by the genome sequencing of the ammonia-oxidizing bacterium Nitrosomonas europaea. Here we have used a phylogenetic approach to study the evolution of the N. europaea RH50 gene, and we show that this gene, probably as a component of an integron cassette, has been transferred to the N. europaea genome by horizontal gene transfer. In addition, by functionally characterizing the Rh50Ne protein and the corresponding knockout mutant, we determined that NeRh50 can mediate ammonium uptake. The RH50Ne gene may thus have replaced functionally the AMT gene, which is missing in the genome of N. europaea and may be regarded as a case of nonorthologous gene displacement. 相似文献
15.
Evolution and functional divergence of monocarboxylate transporter genes in vertebrates 总被引:2,自引:0,他引:2
Monocarboxylate transporters (MCTs) form a gene family with an ancient past. The identification of MCTs (MCHs) from bacteria, protozoa, fungi, invertebrates, as well as vertebrates, but not from plants and virus, allowed illuminating the phylogenetic and evolutionary history of this gene family. The significant expansion of vertebrate MCT genes should have primarily occurred after the divergence of vertebrates and invertebrates, but before the divergence time between ray-finned fish and mammals. The divergence of insect MCTs should have at least occurred in the common ancestor of fruit fly, beetle, and honeybee. Fungi monocarboxylate transporter homologues (MCHs) might evolve independently from an ancient ancestor. The results of functional divergence analysis provided statistical evidences for shifted evolutionary rate and/or changes of amino acid property after gene duplication. The sliding window analysis of the d(N)/d(S) ratio values showed that strong functional constraints must impose on the N- and C-terminal domains of vertebrate MCTs. These corresponding regions may play crucial roles for functionality of MCT proteins. 相似文献
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Horizontal gene transfer in prokaryotes is rampant on short and intermediate evolutionary time scales. It poses a fundamental problem to our ability to reconstruct the evolutionary tree of life. Is it also frequent over long evolutionary distances? To address this question, we analyzed the evolution of 2,091 insertion sequences from all 20 major families in 438 completely sequenced prokaryotic genomes. Specifically, we mapped insertion sequence occurrence on a 16S rDNA tree of the genomes we analyzed, and we also constructed phylogenetic trees of the insertion sequence transposase coding sequences. We found only 30 cases of likely horizontal transfer among distantly related prokaryotic clades. Most of these horizontal transfer events are ancient. Only seven events are recent. Almost all of these transfer events occur between pairs of human pathogens or commensals. If true also for other, non-mobile DNA, the rarity of distant horizontal transfer increases the odds of reliable phylogenetic inference from sequence data. 相似文献
18.
Ehsan Tahrokh Mansour Ebrahimi Mahdi Ebrahimi Fatemeh Zamansani Narjes Rahpeyma Sarvestani Manijeh Mohammadi-Dehcheshmeh Mohammad Reza Ghaemi Esmaeil Ebrahimie 《Genes & genomics.》2011,33(5):565-575
Ammonium is an excellent nitrogen source, and ammonium transfer is a fundamental process in most organisms. Membrane transport of ammonium is the key component of nitrogen metabolism mediated by Ammonium Transporter/Methylamine Permease/Rhesus (AMT/MEP/Rh) protein family. Ammonium transporters play different physiological roles in various organisms. Here, we looked at the protein characteristics of ammonium transporters in different organisms to create a link between protein characteristics and the organism. In order to increase the accuracy and precision of the employed models, for the first time, an attempt was made to cover all structural aspects of ammonium transporters in animals, bacteria, fungi, plants, and human by extracting and calculating 874 protein attributes of primary, secondary, and tertiary structures for each ammonium transporter. Then, various weighting and modeling algorithms were applied to determine how structural protein features change between organisms. Considering a large number of protein attributes made it possible to detect key protein characteristics in the structure of ammonium transporters. The results, for the first time, indicated that His-based features including count/frequency of His and frequency/count of Ile-His were the most significant features generating different types of ammonium transporters within organisms. Within different tested models, the C5.0 model was the most efficient and precise model for discrimination of organism type, based on ammonium transporter sequence, with the precision of 94.85%. The determination of protein characteristics of ammonium transporters in different organisms provides a new vista for understanding the evolution of transporters based on the modulation of protein characteristics and facilitates engineering of new transporters. In our point of view, dissecting a large number of structural protein characteristics through data mining algorithms provides a novel functional strategy for studying evolution and phylogeny. This research will serve as a basis for future studies on engineering novel ammonium transporters. 相似文献
19.
Javelle A Rodríguez-Pastrana BR Jacob C Botton B Brun A André B Marini AM Chalot M 《FEBS letters》2001,505(3):393-398
Heterologous expression of the yeast triple Mep mutant has enabled the first molecular characterization of AMT/MEP family members in an ectomycorrhizal fungus. External hyphae, which play a key role in nitrogen nutrition of trees, are considered as the absorbing structure of the ectomycorrhizal symbiosis and therefore molecular studies on ammonium transport in hyphae are urgently needed. The kinetic properties of AMT2 and AMT3 from Hebeloma cylindrosporum were studied in Saccharomyces cerevisiae. Expression of HcAmts in the yeast triple Mep mutant restored ammonium retention within cells. The HcAmts did not complement the ammonium sensing defect phenotype of Mep2Delta cells during pseudohyphal differentiation. Northern blot analysis in H. cylindrosporum showed that the HcAMTs were up-regulated upon nitrogen deprivation and down-regulated by ammonium. 相似文献