Crystal structures of Magnaporthe oryzae trehalose-6-phosphate synthase (MoTps1) suggest a model for catalytic process of Tps1

2019 ◽  
Vol 476 (21) ◽  
pp. 3227-3240 ◽  
Author(s):  
Shanshan Wang ◽  
Yanxiang Zhao ◽  
Long Yi ◽  
Minghe Shen ◽  
Chao Wang ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Conformational Changes ◽  
Magnaporthe Oryzae ◽  
Structural Information ◽  
Complex Structure ◽  
Critical Role ◽  
Catalytic Process ◽  
Structural Basis ◽  
Salt Bridges ◽  
Terminal Domain

Trehalose-6-phosphate (T6P) synthase (Tps1) catalyzes the formation of T6P from UDP-glucose (UDPG) (or GDPG, etc.) and glucose-6-phosphate (G6P), and structural basis of this process has not been well studied. MoTps1 (Magnaporthe oryzae Tps1) plays a critical role in carbon and nitrogen metabolism, but its structural information is unknown. Here we present the crystal structures of MoTps1 apo, binary (with UDPG) and ternary (with UDPG/G6P or UDP/T6P) complexes. MoTps1 consists of two modified Rossmann-fold domains and a catalytic center in-between. Unlike Escherichia coli OtsA (EcOtsA, the Tps1 of E. coli), MoTps1 exists as a mixture of monomer, dimer, and oligomer in solution. Inter-chain salt bridges, which are not fully conserved in EcOtsA, play primary roles in MoTps1 oligomerization. Binding of UDPG by MoTps1 C-terminal domain modifies the substrate pocket of MoTps1. In the MoTps1 ternary complex structure, UDP and T6P, the products of UDPG and G6P, are detected, and substantial conformational rearrangements of N-terminal domain, including structural reshuffling (β3–β4 loop to α0 helix) and movement of a ‘shift region' towards the catalytic centre, are observed. These conformational changes render MoTps1 to a ‘closed' state compared with its ‘open' state in apo or UDPG complex structures. By solving the EcOtsA apo structure, we confirmed that similar ligand binding induced conformational changes also exist in EcOtsA, although no structural reshuffling involved. Based on our research and previous studies, we present a model for the catalytic process of Tps1. Our research provides novel information on MoTps1, Tps1 family, and structure-based antifungal drug design.

Investigating the dynamic nature of the ABC transporters: ABCB1 and MsbA as examples for the potential synergies of MD theory and EPR applications

2015 ◽  
Vol 43 (5) ◽  
pp. 1023-1032 ◽  
Author(s):  
Thomas Stockner ◽  
Anna Mullen ◽  
Fraser MacMillan
Keyword(s):  
Crystal Structures ◽  
Conformational Changes ◽  
Abc Transporters ◽  
Epr Spectroscopy ◽  
Structural Information ◽  
Dynamic Properties ◽  
Molecular System ◽  
Synergistic Effects ◽  
Md Simulations ◽  
Substrate Spectrum

ABC transporters are primary active transporters found in all kingdoms of life. Human multidrug resistance transporter ABCB1, or P-glycoprotein, has an extremely broad substrate spectrum and confers resistance against chemotherapy drug treatment in cancer cells. The bacterial ABC transporter MsbA is a lipid A flippase and a homolog to the human ABCB1 transporter, with which it partially shares its substrate spectrum. Crystal structures of MsbA and ABCB1 have been solved in multiple conformations, providing a glimpse into the possible conformational changes the transporter could be going through during the transport cycle. Crystal structures are inherently static, while a dynamic picture of the transporter in motion is needed for a complete understanding of transporter function. Molecular dynamics (MD) simulations and electron paramagnetic resonance (EPR) spectroscopy can provide structural information on ABC transporters, but the strength of these two methods lies in the potential to characterise the dynamic regime of these transporters. Information from the two methods is quite complementary. MD simulations provide an all atom dynamic picture of the time evolution of the molecular system, though with a narrow time window. EPR spectroscopy can probe structural, environmental and dynamic properties of the transporter in several time regimes, but only through the attachment sites of an exogenous spin label. In this review the synergistic effects that can be achieved by combining the two methods are highlighted, and a brief methodological background is also presented.

scholarly journals Plasticity at the DNA recognition site of the MeCP2 mCG-binding domain

2019 ◽  
Author(s):  
Ming Lei ◽  
Wolfram Tempel ◽  
Ke Liu ◽  
Jinrong Min
Keyword(s):  
Crystal Structures ◽  
Transcriptional Repression ◽  
Consensus Sequence ◽  
Complex Structure ◽  
Structural Basis ◽  
Structural Studies ◽  
Transcription Activator ◽  
Methylated Dna ◽  
Attachment Region

AbstractMeCP2 is an abundant protein, involved in transcriptional repression by binding to CG and non-CG methylated DNA. However, MeCP2 might also function as a transcription activator as MeCP2 is found bound to sparsely methylated promoters of actively expressed genes. Furthermore, Attachment Region Binding Protein (ARBP), the chicken ortholog of MeCP2, has been reported to bind to Matrix/scaffold attachment regions (MARs/SARs) DNA with an unmethylated 5’-CAC/GTG-3’ consensus sequence. In this study, we investigated how MeCP2 recognizes unmethylated 5’-CAC/GTG-3’ motif containing DNA by binding and structural studies. We found that MeCP2-MBD binds to MARs DNA with a comparable binding affinity to mCG DNA, and the MeCP2-CAC/GTG complex structure revealed that MeCP2 residues R111 and R133 form base-specific interactions with the GTG motif. For comparison, we also determined crystal structures of the MeCP2-MBD bound to mCG and mCAC/GTG DNA, respectively. Together, these crystal structures illustrate the adaptability of the MeCP2-MBD toward the GTG motif as well as the mCG DNA, and also provide structural basis of a biological role of MeCP2 as a transcription activator and its disease implications in Rett syndrome.

A special halide-stabilizing pair and enantioselectivity mechanism of DatA

2014 ◽  
Vol 70 (a1) ◽  
pp. C450-C450
Author(s):  
Lijun Guan ◽  
Hideya Yabuki ◽  
Masahiko Okai ◽  
Jun Ohtsuka ◽  
Masaru Tanokura
Keyword(s):  
Reaction Mechanism ◽  
Crystal Structures ◽  
Structural Information ◽  
Docking Simulation ◽  
Competitive Inhibitor ◽  
Residue Pair ◽  
Structural Basis ◽  
Chiral Compounds ◽  
In The Wild ◽  
Agrobacterium Tumefaciens C58

A novel haloalkane dehalogenase DatA from Agrobacterium tumefaciens C58 belongs to the HLD-II subfamily and hydrolyzes brominated and iodinated compounds, leading to the generation of the corresponding alcohol, a halide ion, and a proton. DatA possesses a unique Asn-Tyr residue pair instead of the Asn-Trp residue pair conserved among the subfamily members, thus the structural basis for its reaction mechanism merits elucidation. In addition, DatA is potentially useful for pharmaceutical and environmental applications, though several crystal structures of HLD-II dehalogenases have been reported so far, the determination of the DatA structure will provide an important contribution to those fields. This work provided insight into the reaction mechanism of DatA via a combination of X-ray crystallographic and computational analysis. The crystal structures of DatA and the Y109W mutant were determined at 1.70 Å [1] and 1.95 Å, respectively. The location of the active site was confirmed by using its novel competitive inhibitor, CHES. The structural information from these two crystal structures and the docking simulation with 1,3-dibromopropane revealed that the replacement of the Asn-Tyr pair with the Asn-Trp pair increases the binding affinity for 1,3-dibromopropane, due to the extra hydrogen bond between Trp109 and halogenated compounds; and that the key residue to bind halogenated substrate is only Asn43 in the wild-type DatA, while those in the Y109W mutant are the Asn-Trp pair. Furthermore, docking simulation using the crystal structures of DatA and some chiral compounds indicated that enantioselectivity of DatA toward brominated alkanes is determined by the large and small spaces around the halogen binding site.

scholarly journals Structure of lipoprotein lipase in complex with GPIHBP1

2019 ◽  
Vol 116 (21) ◽  
pp. 10360-10365 ◽  
Author(s):  
Rishi Arora ◽  
Amitabh V. Nimonkar ◽  
Daniel Baird ◽  
Chunhua Wang ◽  
Chun-Hao Chiu ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Lipoprotein Lipase ◽  
Structural Information ◽  
Density Lipoprotein ◽  
Lipid Binding ◽  
Structural Basis ◽  
Maturation Factor ◽  
Lipase Maturation Factor 1 ◽  
Hydrolysis Of

Lipoprotein lipase (LPL) plays a central role in triglyceride (TG) metabolism. By catalyzing the hydrolysis of TGs present in TG-rich lipoproteins (TRLs), LPL facilitates TG utilization and regulates circulating TG and TRL concentrations. Until very recently, structural information for LPL was limited to homology models, presumably due to the propensity of LPL to unfold and aggregate. By coexpressing LPL with a soluble variant of its accessory protein glycosylphosphatidylinositol-anchored high-density lipoprotein binding protein 1 (GPIHBP1) and with its chaperone protein lipase maturation factor 1 (LMF1), we obtained a stable and homogenous LPL/GPIHBP1 complex that was suitable for structure determination. We report here X-ray crystal structures of human LPL in complex with human GPIHBP1 at 2.5–3.0 Å resolution, including a structure with a novel inhibitor bound to LPL. Binding of the inhibitor resulted in ordering of the LPL lid and lipid-binding regions and thus enabled determination of the first crystal structure of LPL that includes these important regions of the protein. It was assumed for many years that LPL was only active as a homodimer. The structures and additional biochemical data reported here are consistent with a new report that LPL, in complex with GPIHBP1, can be active as a monomeric 1:1 complex. The crystal structures illuminate the structural basis for LPL-mediated TRL lipolysis as well as LPL stabilization and transport by GPIHBP1.

Structure and allosteric regulation of eukaryotic 6-phosphofructokinases

2013 ◽  
Vol 394 (8) ◽  
pp. 977-993 ◽  
Author(s):  
Torsten Schöneberg ◽  
Marco Kloos ◽  
Antje Brüser ◽  
Jürgen Kirchberger ◽  
Norbert Sträter
Keyword(s):  
Crystal Structures ◽  
Structural Information ◽  
Current Knowledge ◽  
Allosteric Regulation ◽  
Site Directed Mutagenesis ◽  
Structural Basis ◽  
Directed Mutagenesis ◽  
Special Focus ◽  
Molecular Architecture ◽  
Key Enzyme

Abstract Although the crystal structures of prokaryotic 6-phosphofructokinase, a key enzyme of glycolysis, have been available for almost 25 years now, structural information about the more complex and highly regulated eukaryotic enzymes is still lacking until now. This review provides an overview of the current knowledge of eukaryotic 6-phosphofructokinase based on recent crystal structures, kinetic analyses and site-directed mutagenesis data with special focus on the molecular architecture and the structural basis of allosteric regulation.

scholarly journals Crystal Structures of Two Coronavirus ADP-Ribose-1″-Monophosphatases and Their Complexes with ADP-Ribose: a Systematic Structural Analysis of the Viral ADRP Domain

2008 ◽  
Vol 83 (2) ◽  
pp. 1083-1092 ◽  
Author(s):  
Yuanyuan Xu ◽  
Le Cong ◽  
Cheng Chen ◽  
Lei Wei ◽  
Qi Zhao ◽  
...  
Keyword(s):  
Structural Analysis ◽  
Crystal Structures ◽  
Structural Information ◽  
Large Family ◽  
Binding Pocket ◽  
Structural Basis ◽  
Active Site Residues ◽  
Replication Process ◽  
Functional Studies ◽  
Group I

ABSTRACT The coronaviruses are a large family of plus-strand RNA viruses that cause a wide variety of diseases both in humans and in other organisms. The coronaviruses are composed of three main lineages and have a complex organization of nonstructural proteins (nsp's). In the coronavirus, nsp3 resides a domain with the macroH2A-like fold and ADP-ribose-1"-monophosphatase (ADRP) activity, which is proposed to play a regulatory role in the replication process. However, the significance of this domain for the coronaviruses is still poorly understood due to the lack of structural information from different lineages. We have determined the crystal structures of two viral ADRP domains, from the group I human coronavirus 229E and the group III avian infectious bronchitis virus, as well as their respective complexes with ADP-ribose. The structures were individually solved to elucidate the structural similarities and differences of the ADRP domains among various coronavirus species. The active-site residues responsible for mediating ADRP activity were found to be highly conserved in terms of both sequence alignment and structural superposition, whereas the substrate binding pocket exhibited variations in structure but not in sequence. Together with data from a previous analysis of the ADRP domain from the group II severe acute respiratory syndrome coronavirus and from other related functional studies of ADRP domains, a systematic structural analysis of the coronavirus ADRP domains was realized for the first time to provide a structural basis for the function of this domain in the coronavirus replication process.

scholarly journals Structural basis of membrane recognition of Toxoplasma gondii vacuole by Irgb6

2021 ◽  
Vol 5 (1) ◽  
pp. e202101149
Author(s):  
Yumiko Saijo-Hamano ◽  
Aalaa Alrahman Sherif ◽  
Ariel Pradipta ◽  
Miwa Sasai ◽  
Naoki Sakai ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Parasitophorous Vacuole ◽  
Membrane Binding ◽  
Structural Basis ◽  
In Silico Docking ◽  
Binding Interface ◽  
Dimerization Interface

The p47 immunity-related GTPase (IRG) Irgb6 plays a pioneering role in host defense against Toxoplasma gondii infection. Irgb6 is recruited to the parasitophorous vacuole membrane (PVM) formed by T. gondii and disrupts it. Despite the importance of this process, the molecular mechanisms accounting for PVM recognition by Irgb6 remain elusive because of lack of structural information on Irgb6. Here we report the crystal structures of mouse Irgb6 in the GTP-bound and nucleotide-free forms. Irgb6 exhibits a similar overall architecture to other IRGs in which GTP binding induces conformational changes in both the dimerization interface and the membrane-binding interface. The membrane-binding interface of Irgb6 assumes a unique conformation, composed of N- and C-terminal helical regions forming a phospholipid binding site. In silico docking of phospholipids further revealed membrane-binding residues that were validated through mutagenesis and cell-based assays. Collectively, these data demonstrate a novel structural basis for Irgb6 to recognize T. gondii PVM in a manner distinct from other IRGs.

scholarly journals Structural basis of membrane recognition of Toxoplasma gondii vacuole by Irgb6

2021 ◽  
Author(s):  
Yumiko Saijo Hamano ◽  
Aalaa Alrahman Sherif ◽  
Ariel Pradipta ◽  
Miwa Sasai ◽  
Naoki Sakai ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Conformational Changes ◽  
Molecular Mechanisms ◽  
Structural Information ◽  
Parasitophorous Vacuole ◽  
Membrane Binding ◽  
Structural Basis ◽  
In Silico Docking ◽  
Binding Interface ◽  
Toxoplasma Gondii Infection

The p47 immunity-related GTPase (IRG) Irgb6 plays a pioneering role in host defense against Toxoplasma gondii infection. It is recruited to the parasitophorous vacuole membrane (PVM) formed by T. gondii and disrupts it. Despite the importance of this process, the molecular mechanisms accounting for PVM recognition by Irgb6 remain elusive due to lack of structural information on Irgb6. Here we report the crystal structures of mouse Irgb6 in the GTP-bound and nucleotide-free forms. Irgb6 exhibits a similar overall architecture to other IRGs in which GTP-binding induces conformational changes in both the dimerization interface and the membrane-binding interface. The membrane-binding interface of Irgb6 assumes a unique conformation, composed of N- and C-terminal helical regions forming a phospholipid binding site. In silico docking of phospholipids further revealed membrane binding residues that were validated through mutagenesis and cell-based assays. Collectively, these data demonstrate a novel structural basis for Irgb6 to recognize T. gondii PVM in a manner distinct from other IRGs.

scholarly journals A CTP-dependent gating mechanism enables ParB spreading on DNA

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Adam SB Jalal ◽  
Ngat T Tran ◽  
Clare EM Stevenson ◽  
Afroze Chimthanawala ◽  
Anjana Badrinarayanan ◽  
...  
Keyword(s):  
Dna Binding ◽  
Crystal Structures ◽  
Chromosome Segregation ◽  
Caulobacter Crescentus ◽  
Nucleation Site ◽  
Structural Basis ◽  
Dna Binding Domain ◽  
Terminal Domain ◽  
Gating Mechanism ◽  
Living Organisms

Proper chromosome segregation is essential in all living organisms. The ParA-ParB-parS system is widely employed for chromosome segregation in bacteria. Previously, we showed that Caulobacter crescentus ParB requires cytidine triphosphate to escape the nucleation site parS and spread by sliding to the neighboring DNA (Jalal et al., 2020). Here, we provide the structural basis for this transition from nucleation to spreading by solving co-crystal structures of a C-terminal domain truncated C. crescentus ParB with parS and with a CTP analog. Nucleating ParB is an open clamp, in which parS is captured at the DNA-binding domain (the DNA-gate). Upon binding CTP, the N-terminal domain (NTD) self-dimerizes to close the NTD-gate of the clamp. The DNA-gate also closes, thus driving parS into a compartment between the DNA-gate and the C-terminal domain. CTP hydrolysis and/or the release of hydrolytic products are likely associated with reopening of the gates to release DNA and recycle ParB. Overall, we suggest a CTP-operated gating mechanism that regulates ParB nucleation, spreading, and recycling.

Molecular Dynamics Simulations of HLA-CW4-Β2M-KIR2DL1 Protein and Homology Modeling of a Complex Associated with Psoriasis Disease (HLA-CW6-Β 2M-KIR2DS1)

2021 ◽  
Author(s):  
Mansour H Almatarneh ◽  
Ahmad M Alqaisi ◽  
Enas K Ibrahim ◽  
Ghada G Kayed ◽  
Joshua W Hollett
Keyword(s):  
Molecular Dynamics ◽  
Conformational Changes ◽  
Complex Structure ◽  
3D Structure ◽  
Md Simulations ◽  
Salt Bridges ◽  
Small Peptide ◽  
Β2 Microglobulin ◽  
Dynamics Simulations ◽  
The Right

Molecular dynamics (MD) simulation was used to study the interactions of two immune proteins of HLA-Cw4-β2m-KIR2DL1 complex with small peptide QYDDAVYKL (nine amino acids) in an aqueous solution. This study aims to gain a detailed information about the conformational changes and the dynamics of the complex. The right parameters and force field for performing the MD simulations that was needed to calibrate the complex structure were determined. The non-bonded interactions (Electrostatic and van der Waals contributions), H-bond formation, and salt bridges between the ligand HLA-Cw4 and the receptor KIR2DL1 were estimated using the obtained MD trajectories. The buried surface area due to binding was calculated to get insight into the causes of specificity of receptor to ligand and explains mutations experiment. The study concluded that β2-microglobulin, one part of the complex, is not directly interacting with the peptide at the groove; therefore, it could be neglected from simulation. Our results showed that β2-microglobulin does not have any significant effect on the dynamics of the 3D-structure of the complex. This project will help in understanding to optimize candidate drug design, a small peptide that disrupts the interaction, for the optimal biological effect.

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