DESIGN, SYNTHESIS AND MOLECULAR DOCKING

STUDY OF
PYRAZINE SUBSTITUTED THIAZOLE DERIVATIVES

Under the supervision of

Mr. P.GOBALA KRISHNAN, M.Pharm.,(Ph.D)

Introduction
Pyrazine Derivatives
as anti tubercular
PYRAZINE
activity:
Pyrazine is a heterocyclic compound contains two nitrogen
atoms in its aromatic ring with molecular formula C4H4N2.
Pyrazine and related heterocyclic compounds play an important
role as intermediates for pharmaceuticals, agricultural chemicals.
O

N
N

NH2

pyrazinamide

Thiazole
Thiazole is a heterocyclic compound featuring both a nitrogen atom
and sulfur atom as part of the aromatic five-membered ring. Thiazole and
related compounds are called 1, 3-azoles (nitrogen and one other
heteroatom in a five-membered ring).
Thiazoles are important class of heterocyclic compounds, found in many
potent biologically active molecules such as Sulfathiazol (antimicrobial
drug), Ritonavir (antiretroviral drug), Abafungin (antifungal drug) with trade
name Abasol cream and Bleomycine and Tiazofurin (antineoplastic drug).

THIAZOLE

Molecular docking
Molecular docking is a method to predict the preferred orientation
of one molecule to a second when bound to each other to form a
stable complex. Computers and programs (software) are used to
predict or simulate the possible reaction (and interactions) between
two molecules based on their 3 dimensional structures.

AutoDock
AutoDock is a Molecular modeling simulation software. Since
2009, it has been open source and is free for non-commercial
usage. It is especially effective for Protein-ligand docking.
AutoDock is a suite of automated docking tools. It is designed to
predict how small molecules, such as substrates or drug
candidates, bind to a receptor of known 3D structure.
Used prior to the experimental screening, the molecular docking,
can be as a powerful computational tool to reduce the labor and
cost of drug development.
AutoDock consists of two main programs:
AutoDock for docking of the ligand to a set of grids describing
the target protein;
AutoGrid for pre-calculating these grids.

AutoDock has applications in:

X-ray crystallography;
structure-based drug design;
lead optimization;
chemical mechanism studies

Usage of AutoDock has contributed to the discovery of

several drugs, including HIV1 integrase inhibitors.

Powerful computational tool to reduce the labor and cost of

drug development.

Receptor or host or lock The "receiving" molecule, most commonly a protein

Ligand or guest or key The complementary partner molecule which binds to the
receptor. Ligands are most often small molecules but could also be another
biopolymer.
Docking Computational simulation of a candidate ligand binding to a receptor.
Binding mode The orientation of the ligand relative to the receptor as well as the
conformation of the ligand and receptor when bound to each other.
Pose A candidate binding mode.
Scoring The process of evaluating a particular pose by counting the number of
favorable intermolecular interactions such as hydrogen bonds and hydrophobic
contacts.
Ranking The process of classifying which ligands are most likely to interact
favorably to a particular receptor based on the predicted free-energy of binding.

Aim and Objective

A brief outline of the research work conducted can be
summarized as follows:
Synthesis of substituted thiazole derivatives.
Synthesis of pyrazine substituted thiazole derivatives.
Selection
of
target
protein
(3-Deoxy-D-arabinoheptulosonate-7-phosphate ) from PDB (PDB ID :2B7O)
Evaluate anti tubercular activity of the synthesized compounds through
molecular docking studies by calculating and comparing the binding
energy , Inhibition constant and hydrogen bonding interaction
between
ligand
and
enzyme
3-Deoxy-D-arabinoheptulosonate-7-phosphate (DAHP) using autodock
software.
Find the high scoring and ranking compound or ligand

Scope And Plan of Work

The incorporation of pyrazine to the
thiazole derivatives to prove how this
combination could influence the biological
activity. A brief outline of the research plan
can be summarised as follows:
Synthesis of substituted thiazole derivatives.
Synthesis of pyrazine substituted thiazole derivatives.
Selection of target protein (3-Deoxy-D-arabinoheptulosonate-7-phosphate ) from PDB (PDB
ID :2B7O)

EXPERIMENT - SCHEME
STEP I: Synthesis of 2-amino thiazole Derivatives
R2

R2
NH2
R1
O

NH2

R3

thiourea

R3

R1

H2N

I(a-j)

Substituted Acetophenone

2-aminothiazole derivatives

STEP II : Synthesis of Pyrazine Substituted thiazole derivative:

R2
Cl

N
N

COOH

+
H2N

R2

R1

N
R3

R1
R3

NH

Substituted Thiazole derivative

6-chloropyrazine-2-carboxylic acid

Cl

II(a-j)
Substituted 2-aminothiazole derivative

List of Substituents:
Compound

R1

IIa

CH3

IIb

OCH3

IIc
IId

NO2

OCH3

R2

R3

---H

---H

---H

---H

---H

---H

OCH3

IIe

---H

OCH3

---H

CF3

IIf

----OH

IIg

CH3
C

OCH3

OCH3

---H

---H

CH3
CH3

IIh

-----Br

---H

---H

IIi

-----Cl

---H

---H

IIj

----OH

---H
OCH3

 General method of Synthesis of substituted

thiazole derivatives (Ia-j) (17)
To a mixture consisting of the appropriate

acetophenone II(a-j) (10 mmol) and thiourea (10

mmol), bromine (20 mmol) was added drop wise
during 30 minutes. The reaction mixture was
heated on a boiling water bath for 10 hours and
water was added to it and again heated until
most of the solid has gone into solution.
The reaction mixture was filtered on hot and the
filtrate was cooled, it was made alkaline with
ammonium hydroxide. The product which
separated was filtered, washed with water and
recrystallized from ethanol

 General method of Synthesis of

pyrazine substituted thiazole
derivatives (IIa-j) (18)
6-chloropyrazine-2-carboxylic acids (50.0
mmol) and thionyl chloride (5.5 mL) was refluxed
in dry toluene (20mL) for about 1h. The crude acyl
chloride dissolved in dry acetone (50 mL) was
added drop wise to a stirred solution of the
corresponding substituted amine (50.0 mmol) in
dry pyridine (50 mL) kept at room temperature.
After the addition was complete, stirring continued
for another 30 min. The reaction mixture was then
poured into cold water (100 mL) and the crude
amide was collected and recrystallized from
aqueous ethanol.

b)

MOLECULAR DOCKING STUDIES-AUTODOCK

PROCEDURE

Preparing the protein :

3-Deoxy-D-arabino-heptulosonate-7phosphate(DAHP):

2B7O (PDB id) , retrieved from the

Brookhaven Protein Data Bank , http://www.rcsb.org/pdb

They were obtained as an identical dimeric structure (two
subunits, chains A and B) , They were simplified to a monomer
by deleting chain B.

Preparing the ligand

Chemsketch file

MDL mol file

Open in Pymolwin

save as PDB file

 Running AutoDock4 :
Docking calculations were performed using Auto dock
software (version 4).
[Make sure the AutoDock executable is in the same
directory as the macromolecule, ligand, GPF, DPF and flex
files
Running: Run Run AutoDock... Launch.

TARGET PROTEIN : 3-Deoxy-D-arabinoheptulosonate-7-phosphate (DAHP)

( PDB ID : 2B7O)

structure and IUPAC name of the synthesized compounds

S.No

Structure with IUPAC name

H
N
N

Cl

6-chloro-N-(4-p-tolylthiazol-2-yl)pyrazine-2-carboxamide

H
N
N

Cl

OCH3

6-chloro-N-(4-(4-methoxyphenyl)thiazol-2-yl)pyrazine-2carboxamide

H
N
N

Cl

NO2

6-chloro-N-(4-(4-nitrophenyl)thiazol-2-yl)pyrazine-2-carboxamide

H
N

N
O
S

Cl

6-chloro-N-(4-(3,4,5-trimethoxyphenyl)thiazol-2-yl)pyrazine-2-carboxamide

H
N
N

N
Cl

S
N

CF3

6-chloro-N-(4-(4-(trifluoromethyl)phenyl)thiazol-2-yl)pyrazine-2-carboxamide

S.No

Structure with IUPAC name

H
N

N
N

Cl

OH

N
O

6-chloro-N-(4-(4-hydroxy-3,5-dimethoxyphenyl)thiazol-2-yl)pyrazine-2-carboxamide

H
N
N

S
C

Cl

N-(4-(4-tert-butylphenyl)thiazol-2-yl)-6-chloropyrazine-2-carboxamide

H
N
N

S
Br

Cl

N-(4-(4-bromophenyl)thiazol-2-yl)-6-chloropyrazine-2-carboxamide

H
N
N

S
Cl

Cl

6-chloro-N-(4-(4-chlorophenyl)thiazol-2-yl)pyrazine-2-carboxamide

10

H
N
N

N
Cl

S
OH
N

6-chloro-N-(4-(4-hydroxy-3-methoxyphenyl)thiazol-2-yl)pyrazine-2-carboxamide

Figure : 1 Docking results of compound IIa with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy =

-10.919 kcal/mole

and hydrogen bond interactions were observed at Tyr533- 2.9 0,

leu 812-3.0 0

Figure : 2 Docking results of compound IIb with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -11.790 kcal/mole
and hydrogen bond interactions were observed at Tyr533- 2.6 0,
Tyr936-3.4 0

Figure : 3 Docking results of compound IIc with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -11.919 kcal/mole
and hydrogen bond interactions were observed at Tyr533- 2.4 0,

Figure : 4 Docking results of compound IId with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -11.869 kcal/mole
and hydrogen bond interactions were observed at Tyr533- 2.4 0,
Leu812- 2.8 0

Figure : 5 Docking results of compound IIe with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -9.472 kcal/mole
and hydrogen bond interactions were observed at Tyr936- 3.0 0,
Leu812- 3.4 0
Tyr936-3.4 0

Figure : 6 Docking results of compound IIf with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -8.827 kcal/mole
and hydrogen bond interactions were observed at Tyr517- 3.0 0,
Leu812- 2.6 0

Figure : 7 Docking results of compound IIg with the active site of the (DAHP). (PDB ID: 2B7O).

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -11.378 kcal/mole
and hydrogen bond interactions were observed at Tyr517- 3.0 0,
Leu812- 2.6 0

Figure : 8 Docking results of compound IIh with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -12.378 kcal/mole
and hydrogen bond interactions were observed at Tyr936- 1.9 0,

Figure : 9 Docking results of compound IIi with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -12.198 kcal/mole
and hydrogen bond interactions were observed at Tyr533- 2.0 & 2.4 0,

Figure : 10 Docking results of compound IIj with the active site of the (DAHP). (PDB ID: 2B7O)

Docking studies are carried out against (DAHP). (PDB ID: 2B7O) :
binding energy = -10.744 kcal/mole
and hydrogen bond interactions were observed at Tyr517- 3.0 & 2.2 0,

SUMMARY AND CONCLUSION

Docking calculations were performed using Auto dock software (version

4).
Docking studies for the synthesized compounds were carried out against

3-Deoxy-D-arabino-heptulosonate-7-phosphate
(DAHP). (PDB ID: 2B7O)
Compounds

carboxamide

N-(4-(4-bromophenyl)thiazol-2-yl)-6-chloropyrazine-2(IIh),

6-chloro-N-(4-(4-chlorophenyl)thiazol-2-

yl)pyrazine-2-carboxamide (IIi) were found to be more potent

anticonvulsant

activity

by

inhibiting

heptulosonate-7-phosphate

3-Deoxy-D-arabino(DAHP).

(PDB

ID:

2B7O) when compared to other synthesized derivatives based upon the

binding energy, and hydrogen bonding formation with active site of
receptor.

Scoring and Ranking of Ligands

Compound Binding
code
energy

Best binding pose (Hydrogen bonding

interaction & distance)

(Kcal/mole)

Rank
(Based on binding
energy)

IIa

-10.919

Tyr533-2.9, leu812-3.0, Asn518-2.7

IIb

-11.790

Tyr533-2.6, 2.2, Tyr936- 3.4,3.5

IIc

-11.919

Tyr533- 2.4

IId

-11.869

Tyr533- 2.4, leu812- 2.8

IIe

-9.4722

Tyr936- 3.0, Leu812-3.4, Tyr517-3.1

IIf

-8.8279

Tyr517-3.0, leu812-2.6

10

IIg

-11.378

Tyr517-3.0, leu812-2.6

IIh

--12.378

Tyr936- 1.9, 3.0

IIi

-12.198

Tyr533- 2.0, 2.4

IIj

-10.744

Tyr517-3.0, 2.2

References
1.

Blumberg, H.M.; Leonard, M.K.; Jasmer, R.M. Update on the treatment of

tuberculosis andlatent tuberculosis infection. JAMA - J. Am. Med. Assoc. 2005, 293,
2776-2784.

2.

Kadam SS, Mahadik KR, Bothara KG. Principles of medicinal chemistry, 6th ed.
Nirali Prakashan, 1999; (I): 169-75.

3.

Satoskar. RS, Bhandarkar SD, Ainapur SS. Pharmacology and pharmacotherapeutics.

18th ed. popular Prakashan Mumbai: 728.

4.

http://en.wikipedia.org/wiki/Tuberculosis

5.

http://en.wikipedia.org/wiki/Tuberculosis treatment.

6.

El-S. T. Ali, A. M. El. Kazak, "Synthesis and antimicrobial activity of some new 1,3thiazoles, 1,3,4-thiadiazoles, 1,2,4-triazoles and 1,3-thiazines incorporating acridine
and 1,2,3,4-tetrahydroacridine moieties". Eur. J. Chem. 1(1), 6-11, 2010.

7.

P. Karegoudar, M. S. Karthikeyan, D. J. Prasad, M. Mahalinga, B. S. Holla, N. S.

Kumari, "Synthesis of some novel 2,4-disubstituted thiazoles as possible antimicrobial
agents". Eur. J. Med. Chem. 43, 261-267, 2008.

P. Vicini, A. Geronkiaki, M. Incerti, F. Zani, J. Dearden, M. Hewitt,"Heteroarylimino-5benzylidene-4-thiazolidinones analogues of 2-thiazolylimino-5-benzylidene-4-thiazolidinoes with

antimicrobial activity: Synthesis and structure-ativity re-lationship".Bioorg. Med. Chem.16, 37143724, 2008.

9. K. M. Basavaraja, B. Somasekhar, S. appalaraju, "synthesis and biological activity of some 2[3-substituted- 2-thione- 1,3,4- thiazole-5-yl]aminobenzothiazoles". Ind. J. Heterocycl. Chem. 18,
69-72, 2008.

T. Karabasanagouda, A.V. Adhikari, D. Ramgopal, G. Para-meshwarappa,"Synthesis of some new 2(4-alkylthiophenoxy)-4- substituted-1,3-thiazoles as possible anti-inflammatory and antimicrobial
agents" Ind. J. Chem. 47B, 144-152, 2008.
M. A. K. Amine, D. E. Abdel Rahman, Y. A. El-Eryani," Synthesis and preliminary evaluation of some
substituted coumarins as anticonvulsant agents". Bioorg. Med. Chem. 16, 5377-5388, 2008.
A. Andreani, M. Rambaldi, A. Leoni, A. Locatelli, R. Bossa, M. Chiericozzi, I. Galatulas, G. Salvatore,"
Synthesis and cardi-otonic activity of imidazo[2,1-b]thiazoles bearing a lactam ring. Eur. J. Med. Chem.
31, 383-387, 1996. B. Jiang, X. H. Gu, "Syntheses and cytotoxicity evaluation of bis (indolyl) thiazole,
bis (indolyl) pyrazinone and bis (indolyl) pyrazine: analogues of cytxic marine bis (indole) alkaloid "
Bioorg. Med. Chem. 8, 363-371, 2000.
B. Jiang, X. H. Gu, "Syntheses and cytotoxicity evaluation of bis (indolyl) thiazole, bis (indolyl)
pyrazinone and bis (indolyl) pyrazine: analogues of cytxic marine bis (indole) alkaloid " Bioorg. Med.
Chem. 8, 363-371, 2000.

THANK YOU