Bogdan Lesyng

Efficacy of 2-halogen substituted d-glucose analogs in blocking glycolysis and killing “hypoxic tumor cells”

AbstractPurpose: Since 2-deoxy-D-glucose (2-DG) is currently in phase I clinical trials to selectively target slow-growing hypoxic tumor cells, 2-halogenated D-glucose analogs were synthesized for improved activity. Given the fact that 2-DG competes with D-glucose for binding to hexokinase, in silico modeling of molecular interactions between hexokinase I and these new analogs was used to determine whether binding energies correlate with biological effects, i.e. inhibition of glycolysis and subsequent toxicity in hypoxic tumor cells. Methods and Results: Using a QSAR-like approach along with a flexible docking strategy, it was determined that the binding affinities of the analogs to hexokinase I decrease as a function of increasing halogen size as follows: 2-fluoro-2-deoxy-D-glucose (2-FG) > 2-chloro-2-deoxy-D-glucose (2-CG) > 2-bromo-2-deoxy-D-glucose (2-BG). Furthermore, D-glucose was found to have the highest affinity followed by 2-FG and 2-DG, respectively. Similarly, flow cytometry and trypan blue exclusion assays showed that the efficacy of the halogenated analogs in preferentially inhibiting growth and killing hypoxic vs. aerobic cells increases as a function of their relative binding affinities. These results correlate with the inhibition of glycolysis as measured by lactate inhibition, i.e. ID50 1 mM for 2-FG, 6 mM for 2-CG and > 6 mM for 2-BG. Moreover, 2-FG was found to be more potent than 2-DG for both glycolytic inhibition and cytotoxicity. Conclusions: Overall, our in vitro results suggest that 2-FG is more potent than 2-DG in killing hypoxic tumor cells, and therefore may be more clinically effective when combined with standard chemotherapeutic protocols.

Ab initio study of proton transfer in [H3N-H-NH3]+ and [H3N-H-OH2]+

Abstract Quantum mechanical ab initio calculations at the MP2/6-31G * level are performed on two proton bound dimer systems, [H 3 N−H−NH 3 ] + and [H 3 N−H−OH 2 ] + . Several calculations using a medium-size polarized basis set were performed as a check of the 6-31G * results. Energies are calculated at heavy-atom separations of 2.25–3.25 A. At fixed monomer separations, H is moved along the intermonomer aixs, thus mapping out the proton transfer potential energy surface. For the ammonia dimer, the energy for displacements of H perpendicular to the N—N axis are also calculated. For the ammonia—water dimer, two different binding geometries for the water molecule are considered. All data are fit to analytical functions. We discuss the effects of squeezing and stretching the donor—acceptor distance on proton transfer.

A comparative study of time dependent quantum mechanical wave packet evolution methods

We present a detailed comparison of the efficiency and accuracy of the second‐ and third‐order split operator methods, a time dependent modified Cayley method, and the Chebychev polynomial expansion method for solving the time dependent Schrodinger equation in the one‐dimensional double well potential energy function. We also examine the efficiency and accuracy of the split operator and modified Cayley methods for the imaginary time propagation.

Structural basis of oncogenic activation caused by point mutations in the kinase domain of the MET proto-oncogene: modeling studies.

Missense mutations in the tyrosine kinase domain of the MET proto‐oncogene occur in selected cases of papillary renal carcinoma. In biochemical and biological assays, these mutations produced constitutive activation of the MET kinase and led to tumor formation in nude mice. Some mutations caused transformation of NIH 3T3 cells. To elucidate the mechanism of ligand‐independent MET kinase activation by point mutations, we constructed several 3D models of the wild‐type and mutated MET catalytic core domains. Analysis of these structures showed that some mutations (e.g., V1110I, Y1248H/D/C, M1268T) directly alter contacts between residues from the activation loop in its inhibitory conformation and those from the main body of the catalytic domain; others (e.g., M1149T, L1213V) increase flexibility at the critical points of the tertiary structure and facilitate subdomain movements. Mutation D1246N plays a role in stabilizing the active form of the enzyme. Mutation M1268T affects the S+1 and S+3 substrate‐binding pockets. Models implicate that although these changes do not compromise the affinity toward the C‐terminal autophosphorylation site of the MET protein, they allow for binding of the substrate for the c‐Abl tyrosine kinase. We provide biochemical data supporting this observation. Mutation L1213V affects the conformation of Tyr1212 in the active form of MET. Several somatic mutations are clustered at the surface of the catalytic domain in close vicinity of the probable location of the MET C‐terminal docking site for cytoplasmic effectors. Proteins 2001;44:32–43.

Dipole moments of 2,4-diketopyrimidines: Part II: Uracil, thymine and their derivatives

Abstract The dipole moments of uracil, thymine and of 29 of their variously substituted derivatives were experimentally determined in dioxane. A vector scheme of calculations was applied to evaluate the effects of alkyl and halogen substituents on the dipole moment of uracil. The dipole moments of a number of diketopyrimidines studied were also calculated quantum mechanically by the CNDO 2 method. General agreement between the experimental and calculated values was obtained. The substituent-induced electron charge redistributions are discussed in terms of σ- and τ-populations.

Structure-based sequence alignment for the β-trefoil subdomain of the clostridial neurotoxin family provides residue level information about the putative ganglioside binding site

Clostridial neurotoxins embrace a family of extremely potent toxins comprised of tetanus toxin (TeNT) and seven different serotypes of botulinum toxin (BoNT/A–G). The β‐trefoil subdomain of the C‐terminal part of the heavy chain (HC), responsible for ganglioside binding, is the most divergent region in clostridial neurotoxins with sequence identity as low as 15%. We re‐examined the alignment between family sequences within this subdomain, since in this region all alignments published to date show obvious inconsistencies with the β‐trefoil fold. The final alignment was obtained by considering the general constraints imposed by this fold, and homology modeling studies based on the TeNT structure. Recently solved structures of BoNT/A confirm the validity of this structure‐based approach. Taking into account biochemical data and crystal structures of TeNT and BoNT/A, we also re‐examined the location of the putative ganglioside binding site and, using the new alignment, characterized this site in other BoNT serotypes.

Barrier to rotation and conformation of the -NR2 group in cytosine and its derivatives. Part II. Experimental and theoretical dipole moments of methylated cytosines☆

The dipole moments of several cytosine, methylaminocytosine and dime-thylaminocytosine derivatives with and without an ortho methyl group were determined experimentally in dioxane and benzene. Calculations of total energies and dipole moments were performed by the CNDO/2 and INDO methods for sp2 and sp3 hybridization of exocyclic nitrogen for different values of rotational angle phiC-N. Comparison of the experimental dipole moments with those calculated for the energy minima suggests that the conformation of the dimethylamino group is not planar and differs from that found in cytosine. 1,5,7-Trimethylcytosine, with the dipole moment of 7 Deby units, was considered to be the model compound which closely reproduces the dipole moment of cytosine.

Adding numbers with DNA

A novel algorithm based on DNA computing for adding binary integer numbers is presented. It requires the unique representation of bits placed in test tubes treated as registers. Amplification step used for the carry operation allows one, in theory, to add numbers with the same quantity of elementary operations, regardless of the number of bits used for representation. New notation proposed in the paper allows for efficient and abstract description of the technical operations on DNA.

MOLECULAR AND ELECTROSTATIC PROPERTIES OF THE N-METHYLATED NUCLEIC ACID BASES BY DENSITY FUNCTIONAL THEORY

Abstract Complete geometry optimizations using a density functional theory (DFT) with the combined Becke3 and LYP functional potentials (B3-LYP) and the conventional ab initio Hartree-Fock (HF) method with the 6-31G(d,p) basis set were carried out for the fundamental tautomeric forms of nucleic acid bases (cytosine, thymine, guanine and adenine) and their derivatives methylated at the N1 (pyrimidines) or N9 (purines) positions. At the HF/6-31G(d,p) geometries, the dipole moments, electronic densities and molecular electrostatic potentials (MEPs) were computed using the HF/6-31G(d,p), MP2(fc)/6-31G(d,p), DFT(B3-LYP)/6-31G(d,p), DFT(B3-LYP)/6-31 + + G(d,p) methods and DFT with inclusion of Becke nonlocal, gradient-corrected exchange energy terms (DFT(NLE) method) with the numerical DNP basis set. The same properties were also computed using the DFT(B3-LYP)/6-31G(d,p) method for the corresponding optimized geometries of the molecules. The charges that reproduce the MEP maps from the ab initio (HF, MP2) and DFT calculations were fitted and compared. The ground state molecular parameters (rotational constants, dipole moments) of the methylated bases are compared with the molecular parameters calculated at the same level for the nonmethylated DNA bases and with available experimental data. The results show that the DFT calculations reproduce well the MP2 results for the MEPs, the ESP charges and the dipole moments of the DNA bases and their N-methylated derivatives.

Quantum-Dynamical Picture of a Multistep Enzymatic Process: Reaction Catalyzed by Phospholipase A 2

A quantum-classical molecular dynamics model (QCMD), applying explicit integration of the time-dependent Schrödinger equation (QD) and Newtonian equations of motion (MD), is presented. The model is capable of describing quantum dynamical processes in complex biomolecular systems. It has been applied in simulations of a multistep catalytic process carried out by phospholipase A(2) in its active site. The process includes quantum-dynamical proton transfer from a water molecule to histidine localized in the active site, followed by a nucleophilic attack of the resulting OH(-) group on a carbonyl carbon atom of a phospholipid substrate, leading to cleavage of an adjacent ester bond. The process has been simulated using a parallel version of the QCMD code. The potential energy function for the active site is computed using an approximate valence bond (AVB) method. The dynamics of the key proton is described either by QD or classical MD. The coupling between the quantum proton and the classical atoms is accomplished via Hellmann-Feynman forces, as well as the time dependence of the potential energy function in the Schrödinger equation (QCMD/AVB model). Analysis of the simulation results with an Advanced Visualization System revealed a correlated rather than a stepwise picture of the enzymatic process. It is shown that an sp(2)--> sp(3) configurational change at the substrate carbonyl carbon is mostly responsible for triggering the activation process.