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International Journal of Molecular... Oct 2018In over 40 years of research on the cellular uptake of auxin it is somewhat chastening that we have elaborated so little on the original kinetic descriptions of auxin... (Review)
Review
In over 40 years of research on the cellular uptake of auxin it is somewhat chastening that we have elaborated so little on the original kinetic descriptions of auxin uptake by plant cells made by Rubery and Sheldrake in 1974. Every aspect of that seminal work has been investigated in detail, and the uptake activity they measured is now known to be attributed to the AUX1/LAX family of permeases. Recent pharmacological studies have defined the substrate specificity of AUX1, biochemical studies have evaluated its permeability to auxin in plant cell membranes, and rigourous kinetic studies have confirmed the affinity of AUX1 for IAA and synthetic auxins. Advances in genome sequencing have provided a rich resource for informatic analysis of the ancestry of AUX1 and the LAX proteins and, along with models of topology, suggest mechanistic links to families of eukaryotic proton co-transporters for which crystal structures have been presented. The insights gained from all the accumulated research reflect the brilliance of Rubery and Sheldrake's early work, but recent biochemical analyses are starting to advance further our understanding of this vitally important family of auxin transport proteins.
Topics: Biological Transport, Active; Cell Membrane; Indoleacetic Acids; Membrane Transport Proteins; Plants
PubMed: 30380696
DOI: 10.3390/ijms19113391 -
Molecular Membrane Biology 2004Our structural knowledge of the major facilitator superfamily (MFS) has dramatically increased in the past year with three structures of proteins from the MFS... (Review)
Review
Our structural knowledge of the major facilitator superfamily (MFS) has dramatically increased in the past year with three structures of proteins from the MFS (oxalate/formate antiporter; lactose/proton symporter and the P(i)/glycerol-3-phosphate antiporter). All three structures revealed 12 transmembrane helices forming two distinct domains and could imply that members of the MFS have preserved both secondary as well as tertiary structural elements during evolution. Lactose permease, a particularly well-studied member of the MFS, has been extensively explored by a number of molecular biological, biochemical and biophysical approaches. In this review, we take a closer look at the structure of LacY and incorporate a wealth of biochemical and biophysical data in order to propose a possible mechanism for lactose/proton symport. In addition, we make some brief comparisons between the structures of LacY and GlpT.
Topics: Animals; Binding Sites; Biological Transport; Humans; Membrane Transport Proteins; Protein Structure, Tertiary; Protons
PubMed: 15371012
DOI: 10.1080/09687680410001716862 -
The Journal of Experimental Biology Nov 1994The lactose permease (lac) of Escherichia coli is a paradigm for membrane transport proteins. Encoded by the lacY gene, the permease has been solubilized, purified to... (Review)
Review
The lactose permease (lac) of Escherichia coli is a paradigm for membrane transport proteins. Encoded by the lacY gene, the permease has been solubilized, purified to homogeneity, reconstituted into phospholipid vesicles and shown to catalyse the coupled translocation of beta-galactosides and H+ with a stoichiometry of unity. Circular dichroism and other spectroscopic approaches demonstrate that the purified permease is about 80% helical. Based on hydropathy analysis of the primary amino-acid sequence, a secondary structure has been proposed in which the protein has 12 hydrophobic domains in alpha-helical conformation that traverse the membrane in zigzag fashion connected by hydrophilic loops. A variety of other approaches are consistent with the model and demonstrate that both the N and C termini are on the inner surface of the membrane, and studies on an extensive series of lac permease/alkaline phosphatase fusion proteins provide exclusive support for the topological predictions of the 12-helix motif. This presentation concentrates on the use of site-directed fluorescence spectroscopy to study structure-function relationships in the permease.
Topics: Amino Acid Sequence; Binding Sites; Cell Membrane; Escherichia coli; Escherichia coli Proteins; Genes, Bacterial; Membrane Transport Proteins; Models, Biological; Models, Structural; Molecular Sequence Data; Monosaccharide Transport Proteins; Protein Structure, Secondary; Symporters
PubMed: 7823021
DOI: 10.1242/jeb.196.1.183 -
Trends in Cell Biology Apr 2010Yeast permeases, that act as transporters for nutrients including amino acids, nucleobases and metals, provide a powerful model system for dissecting the physiological... (Review)
Review
Yeast permeases, that act as transporters for nutrients including amino acids, nucleobases and metals, provide a powerful model system for dissecting the physiological control of membrane protein trafficking. Modification of these transporters by ubiquitin is known to target them for degradation in the vacuole, the degradation organelle of fungi. Recent studies have uncovered the role of specific adaptors for recruiting the Rsp5 ubiquitin ligase to these proteins. In addition, the role of ubiquitin at different trafficking steps including early endocytosis, sorting into the multivesicular body (MVB) pathway and Golgi-to-endosome transit is now becoming clear. In particular, K63-linked ubiquitin chains now emerge as a specific signal for protein sorting into the MVB pathway. A complete view of the ubiquitin code governing yeast permease trafficking might not be far off.
Topics: Animals; Endocytosis; Endosomal Sorting Complexes Required for Transport; Gene Expression Regulation, Enzymologic; Humans; Membrane Proteins; Membrane Transport Proteins; Multivesicular Bodies; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Ubiquitin; Ubiquitin-Protein Ligase Complexes; Vacuoles
PubMed: 20138522
DOI: 10.1016/j.tcb.2010.01.004 -
International Review of Cytology 1992
Review
Topics: Amino Acid Sequence; Biological Transport; Escherichia coli Proteins; Gene Expression; Membrane Transport Proteins; Models, Biological; Molecular Sequence Data; Monosaccharide Transport Proteins; Mutagenesis, Insertional; Mutagenesis, Site-Directed; Protein Structure, Secondary; Protons; Sodium; Symporters
PubMed: 1330966
DOI: 10.1016/s0074-7696(08)62674-1 -
FEBS Letters Nov 2003Membrane transport proteins transduce free energy stored in electrochemical ion gradients into a concentration gradient and are a major class of membrane proteins, many... (Review)
Review
Membrane transport proteins transduce free energy stored in electrochemical ion gradients into a concentration gradient and are a major class of membrane proteins, many of which play important roles in human health and disease. Recently, the X-ray structure of the Escherichia coli lactose permease (LacY), an intensively studied member of a large group of related membrane transport proteins, was solved at 3.5 A. LacY is composed of N- and C-terminal domains, each with six transmembrane helices, symmetrically positioned within the molecule. The structure represents the inward-facing conformation, as evidenced by a large internal hydrophilic cavity open to the cytoplasmic side. The structure with a bound lactose homolog reveals the sugar-binding site in the cavity, and a mechanism for translocation across the membrane is proposed in which the sugar-binding site has alternating accessibility to either side of the membrane.
Topics: Binding Sites; Biological Transport, Active; Carbohydrate Metabolism; Escherichia coli; Escherichia coli Proteins; Membrane Transport Proteins; Models, Molecular; Monosaccharide Transport Proteins; Protein Conformation; Protein Structure, Secondary; Protein Structure, Tertiary; Static Electricity; Symporters; Thermodynamics
PubMed: 14630326
DOI: 10.1016/s0014-5793(03)01087-1 -
BioEssays : News and Reviews in... Dec 1987
Review
Topics: Binding Sites; Biological Transport, Active; Escherichia coli; Escherichia coli Proteins; Membrane Transport Proteins; Monosaccharide Transport Proteins; Mutation; Protein Conformation; Symporters
PubMed: 3325054
DOI: 10.1002/bies.950070608 -
Annual Review of Biophysics and... 1986
Review
Topics: Antibodies, Monoclonal; Biological Transport, Active; Electron Transport; Escherichia coli; Escherichia coli Proteins; Galactosides; Hydrogen-Ion Concentration; Ionophores; Lactose; Membrane Transport Proteins; Membranes; Models, Biological; Monosaccharide Transport Proteins; Mutation; Osmosis; Protein Conformation; Proteolipids; Sodium; Symporters
PubMed: 3521656
DOI: 10.1146/annurev.bb.15.060186.001431 -
Molecular Membrane Biology Jun 2014Förster resonance energy transfer (FRET) is a photophysical process by which a donor (D) molecule in an electronic excited state transfers its excitation energy to a... (Review)
Review
Förster resonance energy transfer (FRET) is a photophysical process by which a donor (D) molecule in an electronic excited state transfers its excitation energy to a second species, the acceptor (A). Since FRET efficiency depends on D-A separation, the measurement of donor fluorescence in presence and absence of the acceptor allows determination of this distance, and therefore FRET has been extensively used as a "spectroscopic ruler". In membranes, interpretation of FRET is more complex, since one D may be surrounded by many A molecules. Such is the case encountered with membrane proteins and lipids in the bilayer. This paper reviews the application of a model built to analyze FRET data between a single tryptophan mutant of the transmembrane protein lactose permease (W151/C154G of LacY), the sugar/H(+) symporter from Escherichia coli, and different pyrene-labeled phospholipids. Several variables of the system with biological implication have been investigated: The selectivity of LacY for different species of phospholipids, the enhancement of the sensitivity of the FRET modeling, and the mutation of a particular aminoacid (D68C) of the protein. The results obtained support: (i) Preference of LacY for phosphatidylethanolamine (PE) over phosphatidylglycerol (PG); (ii) affinity of LacY for fluid (L(α)) phases; and (iii) importance of the aspartic acid in position 68 in the sequence of LacY regarding the interaction with the phospholipid environment. Besides, by exploring the enhancement of the sensitivity by using pure lipid matrices with higher mole fractions of labelled-phospholipid, the dependence on acyl chain composition is unveiled.
Topics: Amino Acid Substitution; Cell Membrane; Escherichia coli; Escherichia coli Proteins; Fluorescence Resonance Energy Transfer; Membrane Transport Proteins; Monosaccharide Transport Proteins; Mutation; Phosphatidylethanolamines; Phosphatidylglycerols; Phospholipids; Symporters
PubMed: 24826799
DOI: 10.3109/09687688.2014.915351 -
Philosophical Transactions of the Royal... Jan 1990Lactose/H+ symport in Escherichia coli is catalysed by a hydrophobic transmembrane protein encoded by the lacY gene that has been purified to homogeneity, reconstituted... (Review)
Review
Lactose/H+ symport in Escherichia coli is catalysed by a hydrophobic transmembrane protein encoded by the lacY gene that has been purified to homogeneity, reconstituted into proteoliposomes and shown to be completely functional as a monomer. Circular dichroic studies and hydropathy profiling of the amino-acid sequence of this 'lac' permease suggest a secondary structure in which the polypeptide consists of 12 hydrophobic segments in alpha-helical conformation that traverse the membrane in zig-zag fashion connected by shorter, hydrophilic domains with most of the charged residues and many of the residues commonly found in beta-turns. Support for certain general aspects of the model has been obtained from other biophysical studies, as well as biochemical, immunological and genetic approaches. Oligonucleotide-directed, site-specific mutagenesis is currently being utilized to probe the structure and function of the permease. Application of the technique provides an indication that Arg302 (putative helix IX), His322 (putative helix X) and Glu325 (putative helix X) may be sufficiently close to hydrogen-bond and that these residues play a critical role in lactose-coupled H+ translocation, possibly as components of a charge-relay type of mechanism. In contrast, Cys residues, which were long thought to play a central role in the mechanism of lactose/H+ symport, do not appear to be involved in either substrate binding or H+ translocation.
Topics: Amino Acid Sequence; Cell Membrane; Escherichia coli; Escherichia coli Proteins; Genes, Bacterial; Membrane Transport Proteins; Molecular Sequence Data; Monosaccharide Transport Proteins; Protein Conformation; Protons; Symporters
PubMed: 1970647
DOI: 10.1098/rstb.1990.0022