scholarly journals Evidence for Pentapeptide-Dependent and Independent CheB Methylesterases

2020 ◽  
Vol 21 (22) ◽  
pp. 8459
Author(s):  
Félix Velando ◽  
José A. Gavira ◽  
Miriam Rico-Jiménez ◽  
Miguel A. Matilla ◽  
Tino Krell
Keyword(s):  
Escherichia Coli ◽  
Molecular Basis ◽  
3D Structure ◽  
Structural Disorder ◽  
Structural Basis ◽  
Signaling Proteins ◽  
Pseudomonas Aeruginosa Pao1 ◽  
Pectobacterium Atrosepticum ◽  
Chemosensory Pathways ◽  
Combined Data

Many bacteria possess multiple chemosensory pathways that are composed of homologous signaling proteins. These pathways appear to be functionally insulated from each other, but little information is available on the corresponding molecular basis. We report here a novel mechanism that contributes to pathway insulation. We show that, of the four CheB paralogs of Pseudomonas aeruginosa PAO1, only CheB2 recognizes a pentapeptide at the C-terminal extension of the McpB (Aer2) chemoreceptor (KD = 93 µM). McpB is the sole chemoreceptor that stimulates the Che2 pathway, and CheB2 is the methylesterase of this pathway. Pectobacterium atrosepticum SCRI1043 has a single CheB, CheB_Pec, and 19 of its 36 chemoreceptors contain a C-terminal pentapeptide. The deletion of cheB_Pec abolished chemotaxis, but, surprisingly, none of the pentapeptides bound to CheB_Pec. To determine the corresponding structural basis, we solved the 3D structure of CheB_Pec. Its structure aligned well with that of the pentapeptide-dependent enzyme from Salmonella enterica. However, no electron density was observed in the CheB_Pec region corresponding to the pentapeptide-binding site in the Escherichia coli CheB. We hypothesize that this structural disorder is associated with the failure to bind pentapeptides. Combined data show that CheB methylesterases can be divided into pentapeptide-dependent and independent enzymes.

scholarly journals Assembly of the B subunit pentamer of Escherichia coli heat-labile enterotoxin. Kinetics and molecular basis of rate-limiting steps in vitro.

1996 ◽  
Vol 271 (43) ◽  
pp. 27188
Author(s):  
Lloyd W. Ruddock ◽  
Jeremy J.F. Coen ◽  
Caroline Cheesman ◽  
Robert B. Freedman ◽  
Timothy R. Hirst
Keyword(s):  
Escherichia Coli ◽  
Molecular Basis ◽  
B Subunit ◽  
Heat Labile ◽  
Rate Limiting

scholarly journals Structural basis for ligand and innate immunity factor uptake by the trypanosome haptoglobin-haemoglobin receptor

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Harriet Lane-Serff ◽  
Paula MacGregor ◽  
Edward D Lowe ◽  
Mark Carrington ◽  
Matthew K Higgins
Keyword(s):  
Innate Immunity ◽  
Trypanosoma Brucei ◽  
Molecular Basis ◽  
Structural Basis ◽  
Β Subunit ◽  
Trypanosome Infection ◽  
Lateral Mobility ◽  
African Trypanosomes ◽  
Distal Half ◽  
Immunity Factor

The haptoglobin-haemoglobin receptor (HpHbR) of African trypanosomes allows acquisition of haem and provides an uptake route for trypanolytic factor-1, a mediator of innate immunity against trypanosome infection. In this study, we report the structure of Trypanosoma brucei HpHbR in complex with human haptoglobin-haemoglobin (HpHb), revealing an elongated ligand-binding site that extends along its membrane distal half. This contacts haptoglobin and the β-subunit of haemoglobin, showing how the receptor selectively binds HpHb over individual components. Lateral mobility of the glycosylphosphatidylinositol-anchored HpHbR, and a ∼50o kink in the receptor, allows two receptors to simultaneously bind one HpHb dimer. Indeed, trypanosomes take up dimeric HpHb at significantly lower concentrations than monomeric HpHb, due to increased ligand avidity that comes from bivalent binding. The structure therefore reveals the molecular basis for ligand and innate immunity factor uptake by trypanosomes and identifies adaptations that allow efficient ligand uptake in the context of the complex trypanosome cell surface.

scholarly journals Structural basis of GTPase-mediated mitochondrial ribosome biogenesis and recycling

2021 ◽  
Author(s):  
Hauke S. Hillen ◽  
Elena Lavdovskaia ◽  
Franziska Nadler ◽  
Elisa Hanitsch ◽  
Andreas Linden ◽  
...  
Keyword(s):  
Molecular Basis ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Large Subunit ◽  
Structural Basis ◽  
Catalytic Core ◽  
Mitochondrial Ribosomes ◽  
Mitochondrial Ribosome ◽  
Translation Systems

Ribosome biogenesis is an essential process that requires auxiliary factors to promote folding and assembly of ribosomal proteins and RNA. In particular, maturation of the peptidyl transferase center (PTC), the catalytic core of the ribosome, is mediated by universally conserved GTPases, but the molecular basis is poorly understood. Here, we define the mechanism of GTPase-driven maturation of the human mitochondrial ribosomal large subunit (mtLSU) using a combination of endogenous complex purification, in vitro reconstitution and cryo-electron microscopy (cryo-EM). Structures of transient native mtLSU assembly intermediates that accumulate in GTPBP6-deficient cells reveal how the biogenesis factors GTPBP5, MTERF4 and NSUN4 facilitate PTC folding. Subsequent addition of recombinant GTPBP6 reconstitutes late mtLSU biogenesis in vitro and shows that GTPBP6 triggers a molecular switch by releasing MTERF4-NSUN4 and GTPBP5 accompanied by the progression to a near-mature PTC state. In addition, cryo-EM analysis of GTPBP6-treated mature mitochondrial ribosomes reveals the structural basis for the dual-role of GTPBP6 in ribosome biogenesis and recycling. Together, these results define the molecular basis of dynamic GTPase-mediated PTC maturation during mitochondrial ribosome biogenesis and provide a framework for understanding step-wise progression of PTC folding as a critical quality control checkpoint in all translation systems.

scholarly journals D-Mannoside FimH Inhibitors as Non-Antibiotic Alternatives for Uropathogenic Escherichia coli

Antibiotics ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1072
Author(s):  
Alfredo Montes-Robledo ◽  
Rosa Baldiris-Avila ◽  
Johan Fabian Galindo
Keyword(s):  
Escherichia Coli ◽  
3D Structure ◽  
Pcr Amplification ◽  
Uropathogenic Escherichia Coli ◽  
Crystallographic Structure ◽  
Type I ◽  
Dynamic Simulations ◽  
Lectin Domain ◽  
In Silico Studies ◽  
Tract Infections

FimH is a type I fimbria of uropathogenic Escherichia coli (UPEC), recognized for its ability to adhere and infect epithelial urinary tissue. Due to its role in the virulence of UPEC, several therapeutic strategies have focused on the study of FimH, including vaccines, mannosides, and molecules that inhibit their assembly. This work has focused on the ability of a set of monosubstituted and disubstituted phenyl mannosides to inhibit FimH. To determine the 3D structure of FimH for our in silico studies, we obtained fifteen sequences by PCR amplification of the fimH gene from 102 UPEC isolates. The fimH sequences in BLAST had a high homology (97–100%) to our UPEC fimH sequences. A search for the three-dimensional crystallographic structure of FimH proteins in the PDB server showed that proteins 4X5P and 4XO9 were found in 10 of the 15 isolates, presenting a 67% influx among our UPEC isolates. We focused on these two proteins to study the stability, free energy, and the interactions with different mannoside ligands. We found that the interactions with the residues of aspartic acid (ASP 54) and glutamine (GLN 133) were significant to the binding stability. The ligands assessed demonstrated high binding affinity and stability with the lectin domain of FimH proteins during the molecular dynamic simulations, based on MM-PBSA analysis. Therefore, our results suggest the potential utility of phenyl mannoside derivatives as FimH inhibitors to mitigate urinary tract infections produced by UPEC; thus, decreasing colonization, disease burden, and the costs of medical care.

Crystal structure of the complex of the interaction domains of Escherichia coli DnaB helicase and DnaC helicase loader: structural basis implying a distortion-accumulation mechanism for the DnaB ring opening caused by DnaC binding

2019 ◽  
Vol 167 (1) ◽  
pp. 1-14
Author(s):  
Koji Nagata ◽  
Akitoshi Okada ◽  
Jun Ohtsuka ◽  
Takatoshi Ohkuri ◽  
Yusuke Akama ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Escherichia Coli ◽  
Molecular Model ◽  
Structural Basis ◽  
Terminal Domain ◽  
Helicase Dnab ◽  
Accumulation Mechanism ◽  
Interaction Domains ◽  
Α Helix ◽  
Available Information

Abstract Loading the bacterial replicative helicase DnaB onto DNA requires a specific loader protein, DnaC/DnaI, which creates the loading-competent state by opening the DnaB hexameric ring. To understand the molecular mechanism by which DnaC/DnaI opens the DnaB ring, we solved 3.1-Å co-crystal structure of the interaction domains of Escherichia coli DnaB–DnaC. The structure reveals that one N-terminal domain (NTD) of DnaC interacts with both the linker helix of a DnaB molecule and the C-terminal domain (CTD) of the adjacent DnaB molecule by forming a three α-helix bundle, which fixes the relative orientation of the two adjacent DnaB CTDs. The importance of the intermolecular interface in the crystal structure was supported by the mutational data of DnaB and DnaC. Based on the crystal structure and other available information on DnaB–DnaC structures, we constructed a molecular model of the hexameric DnaB CTDs bound by six DnaC NTDs. This model suggested that the binding of a DnaC would cause a distortion in the hexameric ring of DnaB. This distortion of the DnaB ring might accumulate by the binding of up to six DnaC molecules, resulting in the DnaB ring to open.

scholarly journals The cryoelectron microscopy structure of the human CDK-activating kinase

2020 ◽  
Vol 117 (37) ◽  
pp. 22849-22857 ◽  
Author(s):  
Basil J. Greber ◽  
Juan M. Perez-Bertoldi ◽  
Kif Lim ◽  
Anthony T. Iavarone ◽  
Daniel B. Toso ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Rational Design ◽  
Control Cell ◽  
3D Structure ◽  
Structural Basis ◽  
Pol Ii ◽  
Insight Into ◽  
Cdk Activating Kinase

The human CDK-activating kinase (CAK), a complex composed of cyclin-dependent kinase (CDK) 7, cyclin H, and MAT1, is a critical regulator of transcription initiation and the cell cycle. It acts by phosphorylating the C-terminal heptapeptide repeat domain of the RNA polymerase II (Pol II) subunit RPB1, which is an important regulatory event in transcription initiation by Pol II, and it phosphorylates the regulatory T-loop of CDKs that control cell cycle progression. Here, we have determined the three-dimensional (3D) structure of the catalytic module of human CAK, revealing the structural basis of its assembly and providing insight into CDK7 activation in this context. The unique third component of the complex, MAT1, substantially extends the interaction interface between CDK7 and cyclin H, explaining its role as a CAK assembly factor, and it forms interactions with the CDK7 T-loop, which may contribute to enhancing CAK activity. We have also determined the structure of the CAK in complex with the covalently bound inhibitor THZ1 in order to provide insight into the binding of inhibitors at the CDK7 active site and to aid in the rational design of therapeutic compounds.

scholarly journals Structural basis for carotenoid cleavage by an archaeal carotenoid dioxygenase

2020 ◽  
Vol 117 (33) ◽  
pp. 19914-19925 ◽  
Author(s):  
Anahita Daruwalla ◽  
Jianye Zhang ◽  
Ho Jun Lee ◽  
Nimesh Khadka ◽  
Erik R. Farquhar ◽  
...  
Keyword(s):  
Active Site ◽  
Molecular Basis ◽  
Structural Basis ◽  
Reaction Intermediates ◽  
Catalytic Activities ◽  
Selective Stabilization ◽  
Carotenoid Cleavage Dioxygenases ◽  
Active Site Cavity ◽  
Binding Cleft ◽  
Shed Light

Apocarotenoids are important signaling molecules generated from carotenoids through the action of carotenoid cleavage dioxygenases (CCDs). These enzymes have a remarkable ability to cleave carotenoids at specific alkene bonds while leaving chemically similar sites within the polyene intact. Although several bacterial and eukaryotic CCDs have been characterized, the long-standing goal of experimentally visualizing a CCD–carotenoid complex at high resolution to explain this exquisite regioselectivity remains unfulfilled. CCD genes are also present in some archaeal genomes, but the encoded enzymes remain uninvestigated. Here, we address this knowledge gap through analysis of a metazoan-like archaeal CCD fromCandidatusNitrosotalea devanaterra (NdCCD).NdCCD was active toward β-apocarotenoids but did not cleave bicyclic carotenoids. It exhibited an unusual regiospecificity, cleaving apocarotenoids solely at the C14′–C13′ alkene bond to produce β-apo-14′-carotenals. The structure ofNdCCD revealed a tapered active site cavity markedly different from the broad active site observed for the retinal-formingSynechocystisapocarotenoid oxygenase (SynACO) but similar to the vertebrate retinoid isomerase RPE65. The structure ofNdCCD in complex with its apocarotenoid product demonstrated that the site of cleavage is defined by interactions along the substrate binding cleft as well as selective stabilization of reaction intermediates at the scissile alkene. These data on the molecular basis of CCD catalysis shed light on the origins of the varied catalytic activities found in metazoan CCDs, opening the possibility of modifying their activity through rational chemical or genetic approaches.

scholarly journals Catechol biosynthesis from glucose in Escherichia coli anthranilate-overproducer strains by heterologous expression of anthranilate 1,2-dioxygenase from Pseudomonas aeruginosa PAO1

2014 ◽  
Vol 13 (1) ◽  
Author(s):  
Víctor E Balderas-Hernández ◽  
Luis G Treviño-Quintanilla ◽  
Georgina Hernández-Chávez ◽  
Alfredo Martinez ◽  
Francisco Bolívar ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Heterologous Expression ◽  
Pseudomonas Aeruginosa Pao1

The structural basis of the high in vivo strength of the rRNA P2 promoter of Escherichia coli

Gene ◽  
1989 ◽  
Vol 78 (1) ◽  
pp. 189-194 ◽  
Author(s):  
Tamas Lukacsovich ◽  
Tamas Gaal ◽  
Pal Venetianer
Keyword(s):  
Escherichia Coli ◽  
Structural Basis ◽  
P2 Promoter
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