cenk-andac-lecture

Computer-Assisted Drug Development (CADD) tools utilized at the Dedicated Computer Network of Turkey

(TR-GRID)

by Cenk (Jenk) A. Andaç

M.S. School of Pharmacy, University of Wisconsin-Madison USA

Ph.D. Candidate School of Pharmacy, Gazi University-Ankara Turkiye

Res. Asst. Department of Chemistry, Dicle University-Diyarbakir Turkiye

[email protected]

The GRID section of this presentation was prepared in collaboration with

Mrs. Feyza Eryol, [email protected]

Mr. Onur Temizsoylu, [email protected]

Mrs. Aslı Zengin, [email protected]

at the National Academic Network and Information (ULAKBIM) of

the Scientific and Technological Research Council of Turkiye (TUBITAK)

Summer School on Molecular Modeling and Drug DesignSeptember 10th-14th, 2008 – The Marmara Hotel , Istanbul

What is GRID?

GRID is a network of high-speed cluster/personal computers communicating via

very fast internet connections to share CPUs and storage area.

Aim :

to boost-up computational power

to increase capacity for data storage

Areas :

- Local GRIDs

small scale GRIDs, institutions, univesities, companies, etc.

- National GRIDs

CERN-Tier0 (Geneva, Switzerland), BNL (Brookhaven, USA),

FZK (Karlsruhe, DE), TR-Grid (Turkiye), etc.

- Global GRIDs (EGEE, OSG, NorduGRID, Tier1, Tier2, etc.)

Types :

- GRID of dedicated clusters EGEE, SEE

- GRID of Super Computer clusters DEISA Infrastructure

- GRID of Desktop computers @Home Projects

TR-10-ULAKBIM TUBITAK-Ankara 2007

93 Nodes

2 X Intel Xeon DUAL-CORE 2.6 GHz CPUs/node

a total of 372 CPUs

4GB RAM / Node

TR-Grid ( National GRID of Turkiye)

Head-Quarter: ULAKBIM-TUBITAK-Ankara

EGEE1 Tbyte64TR-09-ITUIstanbul Technical University

EGEE24 Tbyte+567 Tbyte372TR-10-ULAKBIMTUBITAK ULAKBIM

637 Tbyte

12 Tbyte

1 Tbyte

1 Tbyte

1 Tbyte

24 Tbyte

6 Tbyte

Storage Capacity

Local HPC Users

EGEE

EGEE, SEE-GRID-SCI

EGEE

EGEE, SEE-GRID-SCI

EGEE, SEE-GRID-SCI

Affiliation

240TR-11-ULAKBIMTUBITAK ULAKBIM

1246TOTAL

64TR-07-PAMUKKALEPamukkale University

64TR-05-BOUNBosporus University

64TR-04-ERCIYESErciyes University

312TR-03-METUMiddle-East Technical University

66TR-01-ULAKBIMTUBITAK ULAKBIM

CPUGRID nameLocation

Affiliation of TR-Grid with European Union GRID projects

• EGEE-2 (Enabling Grid for E-SciencE-2, EGEE) (2006-2008)

Aim : Implementation of common projects amongst many nations throughout the world, integration with the existing SEE GRID structures and affiliation with other GRID infrastructures across continents.

Members : Currently registered 224 GRID infrastructures from 49 countries (including TR-Grid -Turkiye)

Capacity : 38 K CPUs and 15 PetaBytes of Data Storage

Budget : ~35 M EURO. 360 K EURO of the total budget is provided by TUBITAK of Turkiye.

Affiliation: EUMED, CERN

Main Projects : LHC-CERN, HERA, Biomed

• SEE-GRID (South Eastern European Grid-enabled e-Infrastructure Development) (2004-2008)

Aim : Implementation of common projects amongst 11 Balkan nations

Members: GRNET (Greece), AMREJ (Yugoslavia), CERN (Switzerland), BAS (Bulgaria), ICI (Romania),

TR-Grid (Turkiye), MTA SZTAKI (Hungary), INIMA (Albania), BIHARNET (Slovenia), MARNET (Macedonia),

RBI (Croatia)

Budget : 2 M EURO. 190 K EURO of the total budget is provided by TUBITAK of Turkiye.

• EUMED (Empowering e-Science across the Mediteranean) (2006-2008)

Aim: Implementation of common projects amongst 14 Mediterranian countries.

Members: CERIST (Algeria), CERN (Switzerland), CNRST (Morocco), Consortium GARR (South-Cyprus),

DANTE (UK), EUN (Greece), HIAST (Syria), INFN (Italy), MRSTDC (Tunisia), RED.ED (Spain),

TR-Grid (Turkiye), University of Malta (Malta).

Budget : 2 M EURO. 79 K EURO is provided by TUBITAK of Turkiye.

Affiliation: EGEE GRID Project,

Tr-GRID projects

International Projects

EGEE projects

Nuclear Physics : LHC-CERN (ATLAS, ALICE, CMS, LHCb), BaBar (Stanford Linear

Accelerator), DESY (HERA-Hamburg electron-proton accelerator), ITER

(Nuclear Fusion)

Astrophysics : ESA-Plack (Satellite Data Processing), MAGIC (Cherenkov Telescope Data Processing),

GeoPhysics : Atmospheric Ozone Monitoring (GOME-ERS-SAR Stalelite Data Analysis), Earthquake

Monitoring, Hydrology (Simulation of Water Movements in Mediterranean Coasts),Weather

Forecast

Biomedical : Medical Imaging, GATE (GEANT4 Application for Tomographic Emission), CDSS (Clinical

Decision Support System), Pharmacokinetics, SiMRI3D (3D MRI Simulator), gPTM3D (Radiology

Analysis), Bronze Standard, SPM (Neuro- Application for Diagnosis in Alzheimer Disease),

Bioinformatics (Protein Sequence Analysis, Electron Microscopy Analysis, Genomiscs), Drug

Development (WISDOM drug development for Malaria and Bird Flu, GridGRAMM-Molecular

Docking, GROCK-Grid Dock)

Computational : GEMS (Grid Enabled Molecular Simulator), ABCtraj (Mono/Di-atomic reaction

Chemistry Analysis) COLOMBUS (ab-initio computations), GAMESS (ab-initio computations, SCF

minimizations, Atomic Charges, QM, etc.)

National Applications

• WP3-> P-GRADE Portal (National GRID Certification Management) ULAKBIM-TUBITAK

• SDA (Kandilli Earthquake Seismic Data Server and Analysis) Bosporus Univ.

• G-PiP (Protein-protein Interaction Prediction Application) Bilkent Univ.& Koç Univ.

• GRIDAE (A Grid-based Framework for Artificial Evolution Applications) METU

• SE4SEE (The Search Engine for South East Europe) Bilkent Univ.

• HuM2S Bosporus Univ.

• University Research Projects (Chemistry, Physics, Life Sciences, Health Sciences)

TR-Grid Projects

CERN-LHC Project @ EGEE

Proton-Proton collision (event) with an energy of 14 TeV in the center of ATLAS

detector.

Raw Data : 15 PetaBytes/year (1 PetaByte=1024 TeraBytes or 10242 GigaBytes

~100.000 CPUs needed to process LHC data every year.

• Available for free to all Turkish University Faculty and Graduate Students.

• New users are required to have a TR-Grid account.

For online applications : or contact TR-Grid at

http://www.grid.org.tr/uyelik [email protected]

TR-Grid TR-Grid users/Connections

Operating System : Scientific Linux Workstation : Linux, Unix, IRIX, Cygwin

Parallel applications : MPI ( MPICH-2 , LAM) Connections : Static IP (University)

Job Scheduling : Torque PBS SSH Secure Shell Connections

(Portable Batch System)

Fortran compilers : gfortran, ifort, pgf PATH/Environment: module avail

Math Kernel Library : Intel MKL module load <name>

module unload <name>

~/.bashrc ~/.cshrc.local

Membership and Use of Tr-GRID

Licenced Software for non-profit academic users

AMBER10 & AMBER_TOOLS1.2 CPMD CHIMERA

CHARMM ABINIT VMD

NAMD VASP SIRIUS

MMTSB_TOOLS RED-III XMGRACE

DOCK 6.2 (UCSF) PWSCF

GAMESS (US)

Commercial Software (License fee is applied per laboratory)

GAUSSIAN

Drug Development Software @TR-Grid


AMBER v10

(Assisted Model Building with Energy Refinement)

http://ambermd.org

MD, QMMM, MM-PBSA, Thermodynamic Integration, Antechamber, Trajectory Analysis, NAB, AMBER Force Fields, LeaP, RESP, etc

DOCK v6.2 (UCSF)

http://dock.compbio.ucsf.edu/DOCK_6/index.htm

Ligand-Receptor Interactions, Conolly Surface, Grid-Based Ligand (library) docking, AMBER scoring.

MMTSB

(Multiscale Modeling Tools for Structural Biology)

http://blue11.bch.msu.edu/mmtsb/Main_Page

Protein/Nucleic Acid Modeling, Aminoacid/Nucleotide Base Mutations, Cluster Analysis

RED-III

(Resp Esp Charge Derive)

http://q4md-forcefieldtools.org/RED/

Requires GAMESS or Gaussian to compute RESP charges for ligands, residues, etc.


$AMBERHOME/bin � 73 AMBER subprograms

links for

gamess, red3, chimera, sirius, xmgrace, mmtsb

AMBER10 Molecular Dynamics

EPtottottottot = Σ Krrrr (r – reqeqeqeq)2222 bondbondsbondsbondsbonds

+ + + + Σ KΘΘΘΘ (Θ – Θeqeqeqeq)2222 angleanglesanglesanglesangles

+ + + + Σ (Vnnnn/2) (1 + cos[nФ – γ]) dihedraldihedraldihedraldihedraldihedral

atomsatomsatomsatoms+ + + + Σ (Aijijijij / R12121212

ijijijij) - (Bijijijij / R6666ijijijij) van der Waals

i i i i ≠≠≠≠ jjjj

atomsatomsatomsatoms+ + + + Σ (qiiii qj j j j / εssss Rijijijij) electrostatic

i i i i ≠≠≠≠ jjjj

[ + [ + [ + [ + EEP extra points (optional) ]]]]atomsatomsatomsatoms

[ [ [ [ ---- ( 1/2) Σ μindindindindiiii . E(o)

iiii polarization (optional) ]]]]i i i i ≠≠≠≠ jjjj


Absolute (single-state) energy terms in MM-PBSA

H = EPtot

EPtot = Hgas + Gsolv

Gsolv = Gel + Gnonel

Gel Poisson Boltzmann

� ε(r)� Ø

(r)= - 4 π . ρ

(r)

Øi = qi [ exp(-κ r) / εw r]

Generalized Born

Gel ≈ GGB = – (1 / 2) (1 – exp {– к .ƒGB} / εw ) Σ ( qiq

j / f GB)

i , j

Gnonel = γ . SASA + b

Stot = Srotational + Stranslational + Svibrational

G = H + T. Stot

Free energy of binding

Receptor + Ligand -> Complex

∆ Gbinding = Gcomplex – [ G receptor + G ligand ]


AMBER10 MM-PB(GB)/SA thermodynamic computations


AMBER MM/PMEMD/QMMM Benchmark Timings @ TR-Grid

PDB ID: 1Z8V for X-Ray Coordinates

MM/PMEMD/QMMM set-up :

Ligand : AM1/GAFF parameters

DNA : HF/6-31G*/ parmbsc0 FF parameters

Complex : Solvated with TIP3 waters in a total

bounding box of 66 Ǻ x 66 Ǻ x 66 Ǻ , neutralized

with counterions (0.28 M Na+)

A total of 13688 atoms

netropsin

MM/PMEMD

2 picoseconds runs

extrapolated to

ns/day & days/100 ns

MD

QMMM

2 picoseconds runs

extrapolated to

ns/day & days/100 ns MD

Ligand: QM/PM3

DNA : MM/Parmbsc0 FF

parameters

Cenk A. Andac, Anooshirvan Miandji, & Ningur Noyanalpan. (2008)

School of Pharmacy, Gazi University-Ankara

Manuscript in preparation


0,00

0,20

0,40

0,60

0,80

1,00

1,20

1,40

1,60

1,80

nan

oseco

nd

s M

D /

day

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 25 30 32 35 40 48

# of CPUs @ Tr-GRID

sander MM Benchmark timings


2n (n = 3,4,5) # of CPUs for high performance by SANDER MD

8 / 16 / 32 CPUs

8 CPUs are prefered for relatively-high speed MD runs

CPU ns MD / day days / 100 ns

1 0,49 204,55

2 0,80 124,65

3 1,00 99,58

4 1,27 79,01

5 1,29 77,80

6 1,45 68,91

7 1,42 70,52

8 1,56 64,02

9 1,54 64,99

10 1,52 65,71

11 1,56 64,13

12 1,60 62,35

13 1,55 64,35

14 1,64 60,80

15 1,64 61,11

16 1,72 58,28

17 1,58 63,19

18 1,60 62,35

19 1,60 62,36

20 1,63 61,17

25 1,60 62,47

30 1,54 64,75

32 1,73 57,81

35 1,55 64,39

40 1,52 65,94

48 1,55 64,68



http://ambermd.org

highest speed for PMEMD : 386 CPUs

speed ratios

MD PMEMD QMMM MD / PMEMD / QMMM

2 0,80 1,25 0,20 1 / 1.56 / 0.25

4 1,27 2,31 0,26 1 / 1.81 / 0.20

8 1,56 3,66 0,29 1 / 2.35 / 0.19

16 1,72 5,13 0,27 1 / 2.98 / 0.16

32 1,73 7,37 0,20 1 / 4.26 / 0.12

48 1,55 8,34 0,19 1 / 5.38 / 0.12

CPUnanoseconds per day

Exemplary CADD/Chemistry Projects @ TR-Grid

(Hacettepe University-Ankara & Dicle University-Diyarbakir)

Cenk Andac, Muge Andac, Adil Denizli (2007). Predicting the Binding Properties of Cibacron Blue F3GA in Affinity Seperation Systems.

International Journal of Biological Macromolecules (Structure, Function and Interactions), 41, 430-438.

Seminar : Cenk Andac. The Third National Affinity Technics. May 2008, Marmaris, Turkiye.

A new manuscript is in preparation.

Development of affinity-columns for efficient removal of HSA out of plasma

Biochemistry Laboratory of Prof. Adil Denizli

at Hacettepe University.

On the right is a Linux workstation directly

connected to Tr-GRID, utilizing AMBER &

DOCK (UCSF)

A CB-F3GA immobilized cryogel column fixed

into a fast protein liquid chromatography (FPLC)

system


Development of affinity-columns for efficient removal of HSA out of plasma

A schematic structure of CB-F3GA immobilized

polyacrylamide cryogels synthesized in Prof. Denizli’s

laboratory at Hacettepe University. This current structure

is yet under development

SEM pictures of the CB-F3GA immobilized cryogels

Exemplary CADD/Chemistry Projects @ TR-GridDevelopment of affinity-columns for efficient removal of HSA out of plasma

• Main objective : Utilization of Cibacron Blue F3GA in the separation of Human

Serum Albumin (HSA) out of human plasma and development of an efficient

affinity chromatography system

• Experimental Approach : Structural analysis on a HSA+CB-F3GA complex

X-ray (Not available)

NMR (Not available)

• Computational Approach : docking and molecular dynamics (MD) studies

�� relevant structures (Protein+CB-F3GA)

�� HSA+CB-F3GA


Bound structures of Cibacron Blue F3GA

Two X-ray structures at the Protein Data Bank

1) Glutathione S-Transferase (GST). An X-Ray structure is available (PDB ID 2OGS)

The GSTs are a family of dimeric enzymes which play a crucial rule in the detoxification of carcinogenic,

mutagenic, toxic and pharmacologically active compounds by conjugating them to the thiol group of the cellular

nucleophile glutathione (GSH, γ-Glu-Cys-Gly)

A.J. Oakley, M. Lo Bello, M. Nuccetelli, A.P. Mazzetti, M.W. Parker, J. Mol. Biol. 291 (1999) 913.

2) CCytosolicytosolic QuinoneQuinone ReductaseReductase (QR)(QR). An X. An X--Ray structure is available (PDB ID 1QRD) Ray structure is available (PDB ID 1QRD) ��

Quinone reductase [NAD(P)H:(quinone acceptor) oxidoreductase is an enzyme that catalyzes

NAD(P)H/FAD(H2)-dependent two-electron reductions of quinones and protects cells against the toxic and

neoplastic (tumor forming) effects of free radicals and reactive oxygen species arising from one-electron

reductions.

R. Li, M.A. Bianchet, P. Talalay, L.M. Amzel, Proc. Natl. Acad. Sci. U.S.A. 92 (1995) 8846.

Solution structure of QR+CB-F3GA+FAD+DQ by Molecular Dynamics

C.A. Andac, M. Andac, A. Denizli. International Journal of Biological Macromolecules (2007), 41, 430-438.

Three binding modes of CBThree binding modes of CB--F3GAF3GA

1) 1) Class I binding mode (CB-F3GA + GST) (PDB ID 2OGS)

2) Class II binding mode (CB-F3GA+QR+FAD+Duroquinone(DQ) (PDB ID 1QRD)

3) Class III binding mode


Binding modes of CB-F3GA

Class I binding mode (CB-F3GA + GST)



Class II binding mode (CB-F3GA+QR+FAD+Duroquinone(DQ)

Cenk Andac, Muge Andac, Adil Denizli (2007). Predicting the Binding Properties of Cibacron Blue F3GA in

Affinity Seperation Systems. International Journal of Biological Macromolecules (Structure, Function and

Interactions), 41, 430-438.

X-Ray structure of Glutathione S-Transferase (GST) dimer (PDB ID 2OGS)



Delphi electrostatic surface map for the final coordinates of 1100 ps molecular dynamics simulation of cytosolic QR in

complex with FAD (green stick view), DQ (orange stick view) and CB-F3GA (atom-color stick view).

Figure was adapted from “Cenk Andac, Muge Andac, Adil Denizli (2007). Predicting the Binding Properties of

Cibacron Blue F3GA in Affinity Seperation Systems. International Journal of Biological Macromolecules (Structure,

Function and Interactions), 41, 430-438.”

•Class III binding mode

..exists in solution structures. All aromatic rings of CB-F3GA are involved in binding


Docking , Molecular Dynamics & AMBER scoring

X-Ray structure of Human Serum Albumin (PDB ID 2BXQ) in complex with myristates, phenylbutazone, indomethacine

J.Ghuman,P.A.Zunszain, I. Petitas,A.A. Bhattacharya, M.Otagiri, S. Curry. JMB 2005, 353, 38.

HSA Binding sitesHSA Binding sites

very-low affinity binding sites

Has been shown to bind BSA at multiple hydrophobic fatty-acid anion binding sites on the surfaceLeatherbarrow RJ, Dean PD (1980). Biochem J. 189(1), 27.

Two well-known high-affinity drug binding sites :SITE-I (warfarin/azapropazone) SITE-II (Benzodiazepins)

Hydrophobic binding site Less-hydrophobic binding sites-------------------------------------------------------------------------------------------------------------------------------

• s-warfarin heme

• r-warfarin ketoprofen

• bilirubin verapamil

• phenylbutazone ibuprofen

• azapropazone indomethacine

• salicylate

• diazepam

• indoxylsulfate

_ W.E. Muller, K.J.Fehske, S.A.C.Schlafer, in : M.M. Reidenberg, S. Erill (Eds.), Drug-

Protein Binding. Praeger, New York, 1986, Chapter 2.

_ G.Sudlow, D.J. Birkett, and D.N. Wade, Mol. Pharmacol. 11:824-32 (1975)

drawbacks for site-I

• presence of a fatty acid in site-I diminishes binding affinty of site-I ligands such as warfarin.

• Less accessibility

• J.M. Ferrer, M.J. Leiton, & A.M.L. Zaton (1998). J. Protein Chem., 17,2,115-119.





Fatty acid binding sites



indomethacin phenylbutazone

CB-F3GA

DOCK experiments – surface structure and spheres



DOCK experiments – docked CB-F3GA



Surface view of HSA in complex with CB-F3GA after 60 ps of MD using AMBER1999 Force Field.



MM-PBSA Thermodynamic computations

• Thermodynamic computations were conducted at 300 K by the mm-pbsa (18) module of AMBER v9 for the complex (HSA+CB-F3GA) exhibiting the following 1:1 binding interaction. Coordinates of the receptor (HSA) and the ligand (CB-F3GA) were extracted from the complex coordinates.

Receptor + Ligand � Complex

• The Absolute free energy (G) for the complex, the receptor, and the ligand was computed in a classical manner as in EQ.1,

G = H – T. S EQ.1

• in which T is the temperature of the system at 300 K. The binding free energy (∆G) of the complex system was computed as in EQ.2

∆G = Gcomp – [ Grec + Glig ] EQ.2

• 10 snapshots were extracted for the coordinates of the solute species (complex, receptor and ligand) at 2 ps time intervals between 50 ps and 60 ps of the trajectory. H and S terms were computed for each snapshots and averaged out of the 10 snapshots to constitute mean absolute H and mean absolute S terms.





�The electrostatic interaction energy in gas phase (∆Eel) between HSA and CB-F3GA is favorable.�The electrostatic contribution to the solvation energy (∆Gel) is disfavorable � The electrostatic contribution to the enthalpy of binding (∆Eel + ∆Gel) is disfavorable (~20 kcal/mol), suggesting that the binding of CB-F3GA displaces all water molecules at the binding interface and energy is consumed to align the ligand and the receptor during the binding process.�Van der Waals interaction energy (∆Evdw) predominantly favors the binding process both in gas and solution phases, yielding a total van der Waals energy (Hwdw) of -84.46 kcal/mol overall, which compensates for the disfavorable effect of electrostatic repulsions, turning the case into a favorable binding state. Internal energy change for the binding process in gas phase (∆Eint) amounts to zero, suggesting that the binding process does not lead to any major conformational violation in the complex.�A total of the entropy terms (translational+rotational+vibrational) makes a disfavorable contribution (+27.24 kcal/mol) to the binding free energy.

Rudimentary MM-PBSA binding energies. All energy values are given in kcal/mol.

| Gas Phase | SGas Phase | Solvation | Total olvation | Total

∆Eel ∆Evdw ∆Eint ∆Hgas ∆Eel ∆Evdw ∆Eint ∆Hgas ∆Gel ∆Gnonel ∆Hgas+∆Gsolv∆Gel ∆Gnonel ∆Hgas+∆Gsolv

H bindingH binding mean σ mean σ mean σ mean σ mean σ mean σ mean σ

EnergiesEnergies -127.40 11.95 -74.62 4.24 0.00 0.0 -202.02 12.48 147.63 8.76 -9.84 0.16 -64.23 7.40

T.∆Strans T.∆Srot T.∆Svib T.∆Stot

S BindingS Binding mean σ mean σ mean σ mean σ

EnergiesEnergies -13.72 0.00 -12.02 0.01 -1.50 11.32 -27.24 11.32

G bindingG binding [∆Hgas + ∆Gsolv] – [T.∆Stot ] = -36.99

energyenergy

Depth of bound CB-F3GA in site II



Spacer lengths



• It is concluded that van der Waals/hydrophobic interactions are the favorable forces

that constitute the enthalpy of binding.

• The binding process of Cibacron Blue F3GA is driven by a favorable enthalpy of

binding.

• The binding process of CB-F3GA onto Serum Albumin adopts a class III binding

mode.

• For a more efficient binding process of immobilized CB-F3GA, an alkanediol spacer

with more than 6 carbons is suggested.

Concluding remarks


Future studies in the laboratories of Denizli et al. at Hacettepe University


Propionic acid derivatives as anti-inflamatuary drugs

• Ibuprofen (PDB ID 2BXG), ketoprofen, Naproxen

• Reported to have binding sites on sites I and II

• Easy to couple them to a spacer with an hydroxyl group.

R-CH(CH3)-COOH + SOCl2 � R-CH(CH3)-COCl

R-CH(CH3)-COCl + HO-spacer-polymer -> R-CH(CH3)-CO-O-spacer-polymer

Docking studies with naproxen

• Naproxen scores 100-1000 times greater affinity for site I of HSA.

• However, accessibility to site I by CB-F3GA immobilized polymers is of great concern.

Alternative drug : Ibuprofen

• Known to bind site II with greater affinity .

Ion-imprinted affinity separation systems for efficient removal of toxic ions out of human

plasma(Hacettepe University & Dicle University)

Serpil Ozkara, Rıdvan Say, Cenk Andac, Adil Denizli (2008). An ion-imprinted monolith for in-vitro removal of iron out of human plasma with

beta thalassemia. Accepted (In Press). Industrial & Engineering Chemistry Research (of the American Chemical Society).

Ahmet Demircelik, Muge Andac, Cenk Andac, Rıdvan Say, Adil Denizli (2008). Molecular recognition-based detoxification of aluminium in

human plasma. Accepted (In Press). Journal of Biomaterials Science: Polymer Edition

Nilgun Candan, Nalan Tuzmen, Muge Andac, Cenk Andac, Rıdvan Say, Adil Denizli. Cadmium removal out of human plasma using ion-

imprinted beads in a magnetic column. Accepted (available online June 8th, 2008). Materials Science & Engineering-C.


The gas-phase structures of (A) [Al3+·(N-

acetylglutamate2-)·(OH-)2·(H2O)2]-1 and (B)

[Al3+·(N-acetylglutamate2-)·(OH-)1·(H2O)3] complex systems, both of which were geometry

optimized using the density functional theory at

the B3LYP/6-31+G(d,p) basis set. H-bonds are

shown with dotted lines.

GAMESS results


Ion-imprinted affinity separation systems for efficient removal of toxic ions out of human plasmaAhmet Demircelik, Muge Andac, Cenk Andac, Rıdvan Say, Adil Denizli (2008). Molecular recognition-based detoxification of aluminium in

human plasma. Accepted (In Press). Journal of Biomaterials Science: Polymer Edition

SEM photographs of (A) PHEMAGA and (B) PHEMAGA-Al3+ beads: Surface

morphology (the upper sides) and bulk structure (the lower sides).

FT-IR

Spectrum of

PHMAGA-Al3+

Beads


RNA Projects

Project-1

Solution structure of the anticodon stem-loop region of a t-RNA with a modified 3-methyl

uridine (3MU) residue at position 33

Cenk Andac, Stephen Scaringe, Ulfert HornemannShool of Pharmacy, University of Wisconsin-Madison, WI USA

Department of Chemistry, Dicle University, Diyarbakir 21280 Turkiye

Dharmacon, Inc., Lafayette, CO, 80026 USA

Methods : RNA synthesis, NMR, AMBER, MMTSB

Manuscript in Preparation

Project-2

Catalytic Small RNAs for protein synthesis

Cenk Andac, Ulfert HornemannShool of Pharmacy, University of Wisconsin-Madison, WI USA

Department of Chemistry, Dicle University, Diyarbakir 21280 Turkiye


Solution structure of the anticodon stem-loop region of a t-RNA with a

modified 3-methyl uridine (3MU) residue at position 32

T-RNAphe (PDB ID: 1EVV)

Synthesis :

2’-(t-butyldimetylsilyl)-

3’-(O-cyanoethyl-N-diisopropylphospho-

amidityl)-5’-(4,4’-dimethoxytrityl)-

3-N-methyluridine

3MU(33)

Jenk Batch Stephen Batch

University of Wisconsin Dharmacon Inc.

Desalting

HPLC

Desalting

NMRJenk RNA

wt_Jenk

1H-NMR spectrum showing two main N-methyl peaks

of 3MU(33) of the 17-mer RNA




Varian NMR data processing

AMBER

51.34%43.21%




AMBER Molecular Dynamics and MMTSB Cluster Analysis

110 ns of MD 2 main clusters were found

Cluster-1 (green)

Cluster-2 (red)

Examplary CADD/Chemistry Projects @ TR-Grid

Two cluster structures of the anticodon stem-loop region of a t-RNA with a


Best structure for cluster-1

at 66.959 ns of MD

(lower potential energy)

Best structure for cluster-2

at 84.774 ns of MD

(higher potential energy)

slow exchange

43.21%

51.34%


MOLECULAR DYNAMICS AND THERMODYNAMICMOLECULAR DYNAMICS AND THERMODYNAMIC PROPERTIES OF HUMAN SIRT2 IN COMPLEX WITH NADPROPERTIES OF HUMAN SIRT2 IN COMPLEX WITH NAD++

Urszula Uciechowska1,Cenk A. Andac3, Ningur Noyanalpan3, Manfred Jung2 and Wolfgang Sippl1

Department of Pharmaceutical Chemistry, Martin-Luther-University Halle-Wittenberg 2Institute for Pharmaceutical Science, Albert-Ludwigs-University Freiburg

3School of Pharmacy, Gazi University-Ankara Turkey

Introduction

NAD+ - dependent histone deacetylases (sirtuins) are enzymes which cleave off the acetyl group from the N-acetylated lysine side-residues of histones and non-histone proteins. Sir2 proteins possess conserved sequences from bacteria to humans and they are able to deacetylate numerous proteins in addition to histones, e.g. α-tubulin, myoD, p53,FOXO [1-3]. The X-ray structures of human and yeast sirtuins as well as the yeast sirtuin in complex with NAD+ have been determined recently. However, the structure of bound NAD+ to human Sirt2 has not yet been determined .

Docking Studies

Figure 1. Obtained NAD conformations

for Sirt2. Docking studies were carried

out using DOCK v6.1 (UCSF) [9]



Root mean-square deviation (RMSD) plots

representing unbound h-Sirt2 (green), bound

h-Sirt2 (red) and bound NAD+ (black) through 9 ns of MD. RMSD plots for bound

and unbound h-Sir2 were referenced to the

X-ray structure coordinates while RMSD for

bound NAD+ was referenced to the docked

coordinates of NAD+.

MM-PBSA thermodynamics results

∆Eele ∆EvdW ∆Gele ∆Gnonel ∆Htot ∆Stot ∆Gcal

Sirt2 NAD -8.60 -47.91 24.89 -6.59 -38.21 -24.98 -13.23

mutation ∆Gmut-bind ∆∆Gbind

1 Gln211Ala -12.52 0.71

2 Arg41Ala -4.08 9.15

3 Asn230Ala -5.13 8.1

4 Ser207Ala -6.64 6.59

5 Glu232Ala -10.39 2.84

6 Lys231Ala -8.91 4.32

7 Gln111Ala -12.90 0.33

8 Val210Ala -6.97 6.26

9 Gln209Ala -6.01 7.22

10 Glu267Ala -12.32 0.91

11 Thr33Ala -11.1 2.13

12 Thr206Ala -6.86 6.37

13 Cys268Ala -9.84 3.39

Alanine Scanning (mutation) Results



ASM binding free energy changes (in kcal/mol) applied on selected residues in h-Sirt2 in complex with NAD+. Residues Arg41, Asn230, Ser207, Val210, Gln209 and Thr206 make the greatest energy contribution to the binding of NAD+.

NAD+ in the binding pocket of h-Sirt2. B.

Residues which make the greatest contributions

to NAD+ binding are shown in green.

∆Gcalc=-13.23 kcal/mol

NAD+ bound to Sirt2 mutated to ARPP. The nicotinamidyl group of NAD+

makes favourable enthalpic contribution to the binding by interaction with Gln209 (green). ARPP is thought to be the waste product of NAD+ that leaves the receptor easily for the next NAD+ to come in and function in the catalytic

site.

∆Gcalc= -5.44 kcal/mol

Thank you for your attention

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