Molecular dynamics simulations of glycosyltransferase LgtC

Šnajdrová, Lenka; Kulhánek, Petr; Koča, Jaroslav

doi:10.1016/j.carres.2003.12.024

Cited by 12 publications

(9 citation statements)

References 25 publications

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“…Large-scale loop rearrangements have been shown to be important for members of families 6, 7 and 8 of GTs. [13][14][15][16][17][18][19] In particular, our results, potentially testable through Cys-Cys crosslinking experiments, 36,37 are consistent with observations from simulation studies of Beta-1,4-Gal-T1 13,14 regarding the functional role of a conserved Trp residue (here: W331), suggesting that the observed loop motions and their coupling to local side-chain conformations may also be relevant for other families of GTs.…”

Section: Discussionsupporting

confidence: 82%

Substrate-induced Conformational Changes and Dynamics of UDP-N-Acetylgalactosamine:Polypeptide N-Acetylgalactosaminyltransferase-2

Milac

Buchete

Fritz

et al. 2007

Journal of Molecular Biology

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O-Glycan biosynthesis is initiated by the transfer of N-acetylgalactosamine (GalNAc) from a nucleotide sugar donor (UDP-GalNAc) to Ser/Thr residues of an acceptor substrate. The detailed transfer mechanism, catalyzed by the UDP-GalNAc polypeptide:N-acetyl-alpha-galactosaminyltransferases (ppGalNAcTs), remains unclear despite structural information available for several isoforms in complex with substrates at various stages along the catalytic pathway. We used all-atom molecular dynamics simulations with explicit solvent and counterions to study the conformational dynamics of ppGalNAcT-2 in several enzymatic states along the catalytic pathway. ppGalNAcT-2 is simulated both in the presence and in the absence of substrates and reaction products to examine the role of conformational changes in ligand binding. In multiple 40-ns-long simulations of more than 600 ns total run time, we studied systems ranging from 45,000 to 95,000 atoms. Our simulations accurately identified dynamically active regions of the protein, as previously revealed by the X-ray structures, and permitted a detailed, atomistic description of the conformational changes of loops near the active site and the characterization of the ensemble of structures adopted by the transferase complex on the transition pathway between the ligand-bound and ligand-free states. In particular, the conformational transition of a functional loop adjacent to the active site from closed (active) to open (inactive) is correlated with the rotameric state of the conserved residue W331. Analysis of water dynamics in the active site revealed that internal water molecules have an important role in enhancing the enzyme flexibility. We also found evidence that charged side chains in the active site rearrange during site opening to facilitate ligand binding. Our results are consistent with the single-displacement transfer mechanism previously proposed for ppGalNAcTs based on X-ray structures and mutagenesis data and provide new evidence for possible functional roles of certain amino acids conserved across several isoforms.

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Section: Discussionsupporting

confidence: 82%

Substrate-induced Conformational Changes and Dynamics of UDP-N-Acetylgalactosamine:Polypeptide N-Acetylgalactosaminyltransferase-2

Milac

Buchete

Fritz

et al. 2007

Journal of Molecular Biology

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“…For those GTs that have crystal structures available, short (<300 ns) all atom or longer (10 µs) course grain molecular dynamic simulations have been performed. These simulations have largely focused upon substrate binding interactions (i.e., LgtC (GT-A), Gpgs (GT-A), and GumK (GT-B)) and in some cases, interactions of the protein with membranes (i.e., alMGS (GT-B) and PglH (GT-B) [ 2 , 27 , 28 , 29 , 30 ]. While multiple enzymes of both the GT-A and GT-B structural classes have been simulated, detailed analysis of the trajectories have only been performed in the GT-A structural scaffolds.…”

Section: Introductionmentioning

confidence: 99%

Conserved Conformational Hierarchy across Functionally Divergent Glycosyltransferases of the GT-B Structural Superfamily as Determined from Microsecond Molecular Dynamics

Ramirez-Mondragon

Nguyen

Milicaj

et al. 2021

IJMS

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It has long been understood that some proteins undergo conformational transitions en route to the Michaelis Complex to allow chemistry. Examination of crystal structures of glycosyltransferase enzymes in the GT-B structural class reveals that the presence of ligand in the active site triggers an open-to-closed conformation transition, necessary for their catalytic functions. Herein, we describe microsecond molecular dynamics simulations of two distantly related glycosyltransferases that are part of the GT-B structural superfamily, HepI and GtfA. Simulations were performed using the open and closed conformations of these unbound proteins, respectively, and we sought to identify the major dynamical modes and communication networks that interconnect the open and closed structures. We provide the first reported evidence within the scope of our simulation parameters that the interconversion between open and closed conformations is a hierarchical multistep process which can be a conserved feature of enzymes of the same structural superfamily. Each of these motions involves of a collection of smaller molecular reorientations distributed across both domains, highlighting the complexities of protein dynamic involved in the interconversion process. Additionally, dynamic cross-correlation analysis was employed to explore the potential effect of distal residues on the catalytic efficiency of HepI. Multiple distal nonionizable residues of the C-terminal domain exhibit motions anticorrelated to positively charged residues in the active site in the N-terminal domain involved in substrate binding. Mutations of these residues resulted in a reduction in negatively correlated motions and an altered enzymatic efficiency that is dominated by lower Km values with kcat effectively unchanged. The findings suggest that residues with opposing conformational motions involved in the opening and closing of the bidomain HepI protein can allosterically alter the population and conformation of the “closed” state, essential to the formation of the Michaelis complex. The stabilization effects of these mutations likely equally influence the energetics of both the ground state and the transition state of the catalytic reaction, leading to the unaltered kcat. Our study provides new insights into the role of conformational dynamics in glycosyltransferase’s function and new modality to modulate enzymatic efficiency.

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“…Similar work has been performed with several GT-As. [26,27] In addition, simulations have been used in tandem with homology modeling to further study GTs when crystal structures are unavailable. [28][29][30][31] Heptosyltransferase I (HepI) is Glycosyltransferase in the GT9 family with a characteristic GT-B fold.…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Ligand Induced Conformational and Dynamical Changes in a GT-B Glycosyltransferase: Molecular Dynamic Simulations of Heptosyltransferase I Apo, Binary and Ternary Complexes

Hassan

Milicaj

et al. 2021

Preprint

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Understanding the dynamical motions and ligand recognition motifs of specific glycosyltransferase enzymes, like Heptosyltransferase I (HepI), is critical to discerning the behavior of other carbohydrate binding enzymes. Prior studies in our lab demonstrated that glycosyltransferases in the GT-B structural class, which are characterized by their connection of two Rossman-like domains by a linker region, have conservation of both structure and dynamical motions, despite low sequence conservation, therefore making discoveries found in HepI transferable to other GT-B enzymes. Through a series of 100 nanosecond Molecular Dynamics simulations of HepI in apo enzyme state, and also in the binary and ternary complexes with the native substrates/products. Ligand free energy analysis allowed determination of an anticipated enzymatic path for ligand binding and release. Principle component, dynamic cross correlation and network analyses of the simulation trajectories revealed that there are not only correlated motions between the N- and C-termini, but also that residues within the N-terminal domain communicate via a path that includes substrate proximal residues of the C-terminal domain. Analysis of structural changes, energetics of substrate/products binding and changes in pKa have elucidated a variety of inter- and intradomain interactions that are critical for catalysis. These data corroborate and allow visualization of previous experimental observations of protein conformational changes of HepI. This study has provided valuable insights into the regions involved in HepI conformational rearrangement upon ligand binding, and are likely to enhance efforts to develop new dynamics disrupting enzyme inhibitors for GT-B structural enzymes in the future.

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Molecular dynamics simulations of glycosyltransferase LgtC

Cited by 12 publications

References 25 publications

Substrate-induced Conformational Changes and Dynamics of UDP-N-Acetylgalactosamine:Polypeptide N-Acetylgalactosaminyltransferase-2

Substrate-induced Conformational Changes and Dynamics of UDP-N-Acetylgalactosamine:Polypeptide N-Acetylgalactosaminyltransferase-2

Conserved Conformational Hierarchy across Functionally Divergent Glycosyltransferases of the GT-B Structural Superfamily as Determined from Microsecond Molecular Dynamics

Ligand Induced Conformational and Dynamical Changes in a GT-B Glycosyltransferase: Molecular Dynamic Simulations of Heptosyltransferase I Apo, Binary and Ternary Complexes

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