RESUMEN
Harnessing genetic diversity in major staple crops through the development of new breeding capabilities is essential to ensure food security1. Here we examined the genetic and phenotypic diversity of the A. E. Watkins landrace collection2 of bread wheat (Triticum aestivum), a major global cereal, by whole-genome re-sequencing of 827 Watkins landraces and 208 modern cultivars and in-depth field evaluation spanning a decade. We found that modern cultivars are derived from two of the seven ancestral groups of wheat and maintain very long-range haplotype integrity. The remaining five groups represent untapped genetic sources, providing access to landrace-specific alleles and haplotypes for breeding. Linkage disequilibrium-based haplotypes and association genetics analyses link Watkins genomes to the thousands of identified high-resolution quantitative trait loci and significant marker-trait associations. Using these structured germplasm, genotyping and informatics resources, we revealed many Watkins-unique beneficial haplotypes that can confer superior traits in modern wheat. Furthermore, we assessed the phenotypic effects of 44,338 Watkins-unique haplotypes, introgressed from 143 prioritized quantitative trait loci in the context of modern cultivars, bridging the gap between landrace diversity and current breeding. This study establishes a framework for systematically utilizing genetic diversity in crop improvement to achieve sustainable food security.
Asunto(s)
Biodiversidad , Productos Agrícolas , Variación Genética , Fenotipo , Fitomejoramiento , Triticum , Alelos , Productos Agrícolas/genética , Introgresión Genética , Variación Genética/genética , Genoma de Planta/genética , Haplotipos/genética , Desequilibrio de Ligamiento/genética , Fitomejoramiento/métodos , Sitios de Carácter Cuantitativo/genética , Triticum/clasificación , Triticum/genética , Secuenciación Completa del Genoma , Filogenia , Estudios de Asociación Genética , Seguridad AlimentariaRESUMEN
ATP-binding cassette (ABC) transporters, although being ubiquitous in biology, often feature a subunit that is limited primarily to bacteria and archaea. This subunit, the substrate-binding protein (SBP), is a key determinant of the substrate specificity and high affinity of ABC uptake systems in these organisms. Most prokaryotes have many SBP-dependent ABC transporters that recognize a broad range of ligands from metal ions to amino acids, sugars and peptides. Herein, we review the structure and function of a number of more unusual SBPs, including an ABC transporter involved in the transport of rare furanose forms of sugars and an SBP that has evolved to specifically recognize the bacterial cell wall-derived murein tripeptide (Mtp). Both these examples illustrate that subtle changes in binding-site architecture, including changes in side chains not directly involved in ligand co-ordination, can result in significant alteration of substrate range in novel and unpredictable ways.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/metabolismo , Adenosina Trifosfato/metabolismo , Bacterias/metabolismo , Proteínas Bacterianas/metabolismo , Transportadoras de Casetes de Unión a ATP/química , Proteínas Bacterianas/química , Sitios de Unión , Evolución Biológica , Modelos Moleculares , Monosacáridos/química , Monosacáridos/metabolismo , Unión Proteica , Estructura Terciaria de ProteínaRESUMEN
The oligopeptide permease (Opp) of Escherichia coli is an ATP-binding cassette transporter that uses the substrate-binding protein (SBP) OppA to bind peptides and deliver them to the membrane components (OppBCDF) for transport. OppA binds conventional peptides 2-5 residues in length regardless of their sequence, but does not facilitate transport of the cell wall component murein tripeptide (Mtp, L-Ala-γ-D-Glu-meso-Dap), which contains a D-amino acid and a γ-peptide linkage. Instead, MppA, a homologous substrate-binding protein, forms a functional transporter with OppBCDF for uptake of this unusual tripeptide. Here we have purified MppA and demonstrated biochemically that it binds Mtp with high affinity (K(D) ⼠250 nM). The crystal structure of MppA in complex with Mtp has revealed that Mtp is bound in a relatively extended conformation with its three carboxylates projecting from one side of the molecule and its two amino groups projecting from the opposite face. Specificity for Mtp is conferred by charge-charge and dipole-charge interactions with ionic and polar residues of MppA. Comparison of the structure of MppA-Mtp with structures of conventional tripeptides bound to OppA, reveals that the peptide ligands superimpose remarkably closely given the profound differences in their structures. Strikingly, the effect of the D-stereochemistry, which projects the side chain of the D-Glu residue at position 2 in the direction of the main chain in a conventional tripeptide, is compensated by the formation of a γ-linkage to the amino group of diaminopimelic acid, mimicking the peptide bond between residues 2 and 3 of a conventional tripeptide.
Asunto(s)
Proteínas Bacterianas/química , Proteínas Portadoras/química , Proteínas de Escherichia coli/química , Escherichia coli/química , Lipoproteínas/química , Proteínas de Transporte de Membrana/química , Peptidoglicano/química , Proteínas Bacterianas/metabolismo , Transporte Biológico , Proteínas Portadoras/metabolismo , Cristalografía por Rayos X , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Lipoproteínas/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Oligopéptidos , Peptidoglicano/metabolismo , Conformación Proteica , EstereoisomerismoRESUMEN
The widespread utilization of sugars by microbes is reflected in the diversity and multiplicity of cellular transporters used to acquire these compounds from the environment. The model bacterium Escherichia coli has numerous transporters that allow it to take up hexoses and pentoses, which recognize the more abundant pyranose forms of these sugars. Here we report the biochemical and structural characterization of a transporter protein YtfQ from E. coli that forms part of an uncharacterized ABC transporter system. Remarkably the crystal structure of this protein, solved to 1.2 A using x-ray crystallography, revealed that YtfQ binds a single molecule of galactofuranose in its ligand binding pocket. Selective binding of galactofuranose over galactopyranose was also observed using NMR methods that determined the form of the sugar released from the protein. The pattern of expression of the ytfQRTyjfF operon encoding this transporter mirrors that of the high affinity galactopyranose transporter of E. coli, suggesting that this bacterium has evolved complementary transporters that enable it to use all the available galactose present during carbon limiting conditions.
Asunto(s)
Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Galactosa/metabolismo , Proteínas de Transporte de Monosacáridos/química , Proteínas de Transporte de Monosacáridos/metabolismo , Pentosas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Transporte Biológico , Cristalografía por Rayos X , Escherichia coli/química , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Cinética , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas de Transporte de Monosacáridos/genética , Operón , Unión Proteica , Alineación de Secuencia , Especificidad por SustratoRESUMEN
Sugar phosphate stress in Escherichia coli is sensed and managed by the transcriptional regulator SgrR and the small RNA (sRNA) SgrS. SgrS is a dual function RNA that performs base pairing-dependent regulation of mRNA targets and encodes a small protein, SgrT. Homologs of SgrR were analyzed for gene synteny and inter-homolog identity to identify those that are likely to be functionally analogous. These 22 SgrR homologs were used to manually locate adjacent sRNAs functionally analogous to SgrS. SgrS homologs shared little sequence identity with E. coli SgrS, but most shared several structural features. The most conserved feature of SgrS homologs was the base pairing region while the most variable feature was the sgrT-coding sequence. Analyses of predicted interactions between SgrS:ptsG mRNA pairs in different organisms revealed interesting differences in the patterns of base pairing interactions. RNA pairs with more interrupted regions of complementarity had a higher proportion of G:C base pairs than those with longer contiguous stretches of complementarity. The identification of this set of homologous sRNAs and their targets sets the stage for future studies to further elucidate the molecular requirements for regulation by SgrS.
Asunto(s)
Proteínas Bacterianas/química , ARN Bacteriano/química , Factores de Transcripción/química , Secuencia de Aminoácidos , Proteínas Bacterianas/clasificación , Proteínas Bacterianas/genética , Emparejamiento Base , Proteínas de Escherichia coli/química , Proteínas de Escherichia coli/clasificación , Gammaproteobacteria/genética , Datos de Secuencia Molecular , Filogenia , ARN Mensajero/química , Homología de Secuencia de Aminoácido , Sintenía , Factores de Transcripción/clasificación , Factores de Transcripción/genéticaRESUMEN
SUMMARY: EchoLOCATION is a database that provides a comprehensive analysis of the subcellular locations of Escherichia coli K-12 proteins. Locations are predicted by integrating data from a range of publicly available algorithms combined with extensive curation of experimental literature. The data can be searched in a variety of ways and can generate lists of subcellular proteomes for analysis. Experimental evidence supports the locations of over 500 envelope proteins (periplasm, inner and outer membrane). From analysis of disagreements between in silico predictions and experimental data, we provide an analysis of protein types where subcellular prediction algorithms are currently not accurate.
Asunto(s)
Algoritmos , Bases de Datos de Proteínas , Proteínas de Escherichia coli/análisis , Escherichia coli/metabolismo , Proteínas de la Membrana Bacteriana Externa/análisis , Proteínas de la Membrana Bacteriana Externa/química , Proteínas de Escherichia coli/química , Modelos Biológicos , Periplasma/química , Proteoma/análisis , Proteómica/métodosRESUMEN
Campylobacter jejuni is a Gram-negative food-borne pathogen associated with gastroenteritis in humans as well as cases of the autoimmune disease Guillain-Barré syndrome. C. jejuni is asaccharolytic because it lacks an active glycolytic pathway for the use of sugars as a carbon source. This suggests an increased reliance on amino acids as nutrients and indeed the genome sequence of this organism indicates the presence of a number of amino acid uptake systems. Cj0982, also known as CjaA, is a putative extracytoplasmic solute receptor for one such uptake system as well as a major surface antigen and vaccine candidate. The crystal structure of Cj0982 reveals a two-domain protein with density in the enclosed cavity between the domains that clearly defines the presence of a bound cysteine ligand. Fluorescence titration experiments were used to demonstrate that Cj0982 binds cysteine tightly and specifically with a K(d) of approximately 10(-7) M consistent with a role as a receptor for a high-affinity transporter. These data imply that Cj0982 is the binding protein component of an ABC-type cysteine transporter system and that cysteine uptake is important in the physiology of C. jejuni.
Asunto(s)
Transportadoras de Casetes de Unión a ATP/química , Transportadoras de Casetes de Unión a ATP/metabolismo , Sistemas de Transporte de Aminoácidos Neutros/química , Sistemas de Transporte de Aminoácidos Neutros/metabolismo , Campylobacter jejuni/química , Cisteína/metabolismo , Transportadoras de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/aislamiento & purificación , Secuencia de Aminoácidos , Sistemas de Transporte de Aminoácidos Neutros/genética , Sistemas de Transporte de Aminoácidos Neutros/aislamiento & purificación , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Sitios de Unión , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Cristalografía por Rayos X , Cisteína/química , Citoplasma/metabolismo , Modelos Moleculares , Datos de Secuencia Molecular , Conformación Proteica , Estructura Terciaria de Proteína , Homología de Secuencia de Aminoácido , Espectrometría de Fluorescencia , Tirosina/química , Tirosina/metabolismoRESUMEN
EchoBASE (http://www.ecoli-york.org) is a relational database designed to contain and manipulate information from post-genomic experiments using the model bacterium Escherichia coli K-12. Its aim is to collate information from a wide range of sources to provide clues to the functions of the approximately 1500 gene products that have no confirmed cellular function. The database is built on an enhanced annotation of the updated genome sequence of strain MG1655 and the association of experimental data with the E.coli genes and their products. Experiments that can be held within EchoBASE include proteomics studies, microarray data, protein-protein interaction data, structural data and bioinformatics studies. EchoBASE also contains annotated information on 'orphan' enzyme activities from this microbe to aid characterization of the proteins that catalyse these elusive biochemical reactions.