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VOLUME 2 | NUMBER 2 | JUN | 2012 |82

Available online at www.ijntps.org | ISSN: 2277 – 2782 INTERNATIONAL JOURNAL OF NOVEL TRENDS IN PHARMACEUTICAL SCIENCES

Molecular Mechanics And Drug Docking Studies On Klebsheilla Pneumonia

(Multidrug Efflux System Protein) Using Insilico Protocols

Hepzibah. W1*, John William

1 S and Balaji. M

2

1Department of Advanced Zoology and Biotechnology, (SECNARM), Loyola College, Chennai-34. Tamilnadu. 2Director, Akshaya Neuroinformatics Research Center, Chennai-5. Tamilnadu

INTRODUCTION

K. pneumoniae is a gram negative bacterium. It is

facultative anaerobic. It is rod-shaped and measures 2 µm

by 0.5 µm. In 1882, Friedlander C. Uber first

discovered Klebsiella to be a pathogen that caused

pneumonia (8). Many hospital cases around the world have

been linked to K. pneumoniae. Therefore, more studies of

the strains were important and performed. The bacterium

was isolated and sequenced from a patient in 2004. K.

pneumoniae is commonly found in the gastrointestinal tract

and hands of hospital personnel (3). The reason for its

pathogenicity is the thick capsule layer surrounding the

bacterium. It is 160 nm thick of fine fibers that protrudes

out from the outer membrane at right angles(6) (5). Another

site on the human body that this bacteria can be found is

the nasopharynx. Its habitat is not limited to humans but is

ubiquitous to the ecological environment. This includes

surface water, sewage, and soil (4).

The complete genome was determined in 2006 at

the Genome Sequencing Center at Washington University in

St. Louis. The genome was named Klebsiella

pneumoniae subsp. pneumoniae MGH 78578. It includes

one chromosome of 5.3 Mbp. The GC content is 57%. There

are five plasmids, pKPN3, pKPN4, pKPN5, pKPN6, and

pKPN7. Respectively, each plasmid length is 0.18 Mbp, 0.11

Mbp, 0.089 Mbp, 0.0043 Mbp, and 0.0035 Mbp. The DNA is

circular. The sequence of K. pneumoniae genome was found

to be closely related that of Escherichia coli K-12 (1).

K. pneumoniae contains a capsule around its cell.

Known as K antigen, it is to protect the bacteria from

phagocytosis. "K. pneumoniae strains of serotypes 01:Kl,

01:K10, and O1:K16, which have only the K antigen exposed

at the cell surface, resist complement-mediated killing by

impeding complement activation. It is also clear that

purified capsular polysaccharides (K antigen) from nine

different serotypes (able or unable to mask the LPS) were

unable to activate complement ." In 1992, K.

pneumoniae could be determined apart from other species

ofKlebsiella. Two oligonucleotide probes and the

hydroxylated fatty acid C14:0-2OH are distinctive of K.

pneumoniae (2). CPS and LPS O side chain are two of the

most important virulence factors of K. pneumoniae (7)

Research Materials

• NCBI ,PUBMED, PUBMED CENTRAL,OMIM,MICROBIAL

GENOME DATABASES,CPH MODEL 2.0 SERVER, Discovery

studio software, MOLSOFT ,RAPPER,PATCHDOCK SERVER

Literature collection: The gene information of Human

Pathogens Klebsheilla pneumonia was collected through

Article Info

Article history

Received 20 May 2012

Revised 25 May 2012

Accepted 19 Jun 2012

Available online 30 Jun 2012

Keywords

Micro-organisms , Protein Modeling

and Drug Docking

Abstract

Klebsiella pneumonia is a necrotizing process with a predilection for debilitated

people. It has a high mortality rate of approximately 50% even with antimicrobial therapy.

The mortality rate approaches 100% for persons with alcoholism and bacteremia.

Klebsiella bacteremia and sepsis produce clinical manifestations similar to those caused by

other gram-negative enteric organisms. Morbidity and mortality rates are comparable to

those for other gram-negative organisms that cause sepsis and septic shock. In neonatal

units, outbreaks caused by ESBL-producing strains present a more serious problem and

may be associated with increased mortality .The primary aim of the present research

investigation is to find out whether the best plant extract compound inhibits the potential

Human pathogens using in-silico methods.

RESEARCH ARTICLE

▼ To whom correspondence should be addressed:

H Hepzibah. W

E-Mail address: [email protected]

Hepzibah et al., Molecular Mechanics and Drug Docking Studies On

System Protein) using Insilico Protocols

VOLUME 2 | NUMBER 2 | JUN | 2012

NCBI –PMC (Pub Med central database).

Gene identification: The genome information and gene

functional studies are identified using Comparative genome

database – Microbial genome databases (MBGDB).

Sequence retrieval system: The best identified potential

microbial proteins sequences were collected through NCBI

database in FASTA format in order to perform molecular

modelling.

Protein modeling: The protein sequences are applied into

CPH SERVER (An Automated Homology Modelling Server)

in order to convert the sequence into 3 D structure.

RESULTS AND DISCUSSION

>gi|238547966|dbj|BAH64317.1| multidrug efflux system protein [Klebsiella pneumoniae subsp.

pneumoniae NTUH-K2044]

MTDLPASVRWQLWIVAFGFFMQSLDTTIVNTALPSMAKSLGESPLHMHMIIVSYVLTVAVMLPASGWLAD

RVGVRNIFFTAIVLFTAGSLFCAQASTLDQLVMARVLQGVGGAMMVPVGRLTVMKIVPRDQYMAAMTFVT

LPGQVGPLLGPALGGVLVEYASWHWIFLINIPVGIVGAIATLCLMPNYTMQTRRFDLSGFLLLAAGMATL

TLALDGQKGLGISPAWLAGLVAVGLCALLLYLWHARGNARALFSLNLFRNRTFSLGLGGSFAGRIGSGML

PFMTPVFLQIGLGFSPFHAGLMMIPMVLGSMGMKRIVVQVVNRFGYRRVLVASTLGLAAVSLLFMFSALA

GWYYVLPLVLFLQGMINASRFSSMNTLTLKDLPDDLASSGNS

MSLDAASTHQVFLYTYLSMAAIIALPALIFSRVPDDVGSNIVLRRRNRSGS

nd Drug Docking Studies On Klebsheilla Pneumonia (Mu

The genome information and gene

ed using Comparative genome

Microbial genome databases (MBGDB).

The best identified potential

microbial proteins sequences were collected through NCBI

database in FASTA format in order to perform molecular

The protein sequences are applied into

CPH SERVER (An Automated Homology Modelling Server)

in order to convert the sequence into 3 D structure.

Molecular visualization tools:

of proteins files are viewed with the

STUDIO SOFTWARE to find out the structural and functional

regions of the 3D structure of the pathogen proteins.

Molecular drug docking:

performed using PATCHDOCK server

docking software) in order to identify the binding affinities

of the pathogen proteins and to identify the (

Quercitrin, Chlorogenic acid, Hyperoside and Rutoside

Chemical Compounds.

Figure: 1Chromosome map







GWYYVLPLVLFLQGMINASRFSSMNTLTLKDLPDDLASSGNSLLSMVMQLSMSIGVTIAGLLLGLYGQQH

MSLDAASTHQVFLYTYLSMAAIIALPALIFSRVPDDVGSNIVLRRRNRSGS

(Multidrug Efflux

|83

Molecular visualization tools: The modelled 3 D structure

of proteins files are viewed with the help of DISCOVERY

STUDIO SOFTWARE to find out the structural and functional

regions of the 3D structure of the pathogen proteins.

Molecular drug docking: The drug docking studies were

performed using PATCHDOCK server- automated molecular

in order to identify the binding affinities

of the pathogen proteins and to identify the (Caffeic acid,

acid, Hyperoside and Rutoside )







LLSMVMQLSMSIGVTIAGLLLGLYGQQH

Hepzibah et al., Molecular Mechanics and Drug Docking Studies On Klebsheilla Pneumonia (Multidrug Efflux


84 | VOLUME 2 | NUMBER 2 | JUN | 2012

Figure: 2 3 Dimensional structure of (Klebsheilla pneumonia (multidrug efflux system protein).

Discovery studio software – Atom model

Figure: 3 3 Dimensional structure of Klebsheilla pneumonia (multidrug efflux system protein).

Discovery studio software – Solid Ribbon model




Figure : 4 Molecular docking Klebsheilla pneumonia (multidrug efflux system protein) and

Caffeic acid

Figure: 5 Molecular docking Klebsheilla pneumonia (multidrug efflux system protein) and

Chlorogenic acid




Figure: 6 Molecular docking Klebsheilla pneumonia (multidrug efflux system protein) and Rutin

Figure: 7Molecular docking Klebsheilla pneumonia (multidrug efflux system protein) and

Quercitrin




Figure: 8 Molecular docking Klebsheilla pneumonia (multidrug efflux system protein) and

Hyperoside

Table: 1

Pathogens Receptor

protein

Caffeic acid Quercitrin Chlorogenic

acid

Hyperoside Rutoside

Klebsheilla

pneumonia

(multidrug efflux

system protein )

-156.98 -309.09 -285.93 -328.08 -429.00

Discussion

Multiple drug resistance or Multidrug resistance is a

condition enabling a disease-causing organism to resist

distinct drugs or chemicals of a wide variety of structure

and function targeted at eradicating the organism.

Organisms that display multidrug resistance can be

pathologic cells, including bacterial and neoplastic (tumor)

cells.

Until recently, research and development (R&D)

efforts have provided new drugs in time to treat bacteria

that became resistant to older antibiotics. That is no longer

the case. The potential crisis at hand is the result of a

marked decrease in industry R&D, and the increasing

prevalence of resistant bacteria. Infectious disease

physicians are alarmed by the prospect that effective

antibiotics may not be available to treat seriously ill patients

in the near future.

As bacterial antibiotic resistance continues to

exhaust the supply of effective antibiotics, a global public

health disaster appears likely. Poor financial investment in

antibiotic research has exacerbated the situation. A call to

arms raised by several prestigious scientific organizations a

few years ago rallied the scientific community, and now the

scope of antibacterial research has broadened considerably

Quercitrin is a glycoside formed from the flavonoid

quercetin and the deoxy sugar rhamnose. It is a constituent

of the dye quercitron. It can be found in Tartary buckwheat

(Fagopyrum tataricum)

Caffeic acid is a hydroxycinnamic acid, a naturally

occurring organic compound. This yellow solid consists of

both phenolic and acrylic functional groups. It is found in all

plants because it is a key intermediate in the biosynthesis of

lignin, one of the principal sources of biomass. Caffeic acid

has been shown to inhibit carcinogenesis, although other

experiments show possible carcinogenic effects. It is also




known as an antioxidant in vitro and also in vivo. Caffeic

acid also shows immunomodulatory and antiinflammtory

activity. 4

Chlorogenic acid is a hydroxycinnamic acid, a

member of a family of naturally occurring organic

compounds. These are esters of polyphenolic caffeic acid

and cyclitol (-)-quinic acid (Clifford, M. N et al., 2003). It is

an important biosynthetic intermediate ( Wout Boerjan , et

al 2003) .It also is one of the phenols found in coffee,

bamboo Phyllostachys edulis (Kweon,Mee-Hyang et al.,

2001), as well as many other plants(Clifford, M. N. (2003)

).This compound, long known as an antioxidant, also slows

the release of glucose into the bloodstream after a

meal(Johnston, K. L et al ., 2003).

Hyperoside is a chemical compound. It is the 3-O-

galactoside of quercetin. It is a medicinally active

compound that can be isolated from Drosera rotundifolia,

from the Stachys plant, from Prunella vulgaris, from Rumex

acetosella, Cuscuta chinensis seeds, from St John's wort and

from Camptotheca acuminata (Shiyou Li , et al., 2005). In

Rheum nobile and R. rhaponticum, it serves as a UV blocker

found in the bracts.It can have a protective antioxidant

effect on cultured PC12 cells. (Zhiyong Liu , et al., 2005).

Rutin, also called rutoside, quercetin-3-O-rutinoside

and sophorin, is a citrus flavonoid glycoside found in

buckwheat, (Kreft S et al., 1999). It can combine with

cations, supplying nutrients from the soil to the cells in

plants. In humans, it attaches to the iron ion Fe2+,

preventing it from binding to hydrogen peroxide, which

would otherwise create a highly-reactive free radical that

may damage cells. It is also an antioxidant.

Furthermore, it has been shown to inhibit in vitro

the vascular endothelial growth factor (Luo; et al., 2008) .in

subtoxic concentrations, so acts as an inhibitor of

angiogenesis. This finding can be potentially relevant for

the control of some cancers (table 2).

Docking is frequently used to predict the binding

orientation of small molecule drug candidates to their

protein targets in order to in turn predict the affinity and

activity of the small molecule. Hence docking plays an

important role in the rational design of drugs. 5

The extracted chemical compound structure and

the predicted three-dimensional structure of Human

pathogen protein were docked using PATCHDOCK server.

Molecular mechanics methods may also be used to provide

semi-quantitative prediction of the binding affinity.

The table (3and 4) in fig (1 and 2) show the binding

affinities between the Ligand and the receptor. The Rutin

shows the high binding affinities (-429.00 /Klebsheilla

pneumoniae) .An overall affinity shows that Hypersoide and

Rutin are the potential inhibitors of the target (Klebsheilla

pneumonia )pathogen proteins.

Conclusion

Antibiotic resistance is an important tool for genetic

engineering. We focus on how the naturally occurring

chemicals inhibit the Multi-drug Resistance (MDR) Human

pathogens (Klebsheilla pneumonia,) protein. Thus, we

identified that the chemical compounds in the plant

products are best agents to reduce the problems associated

to urinary tract infections. So we conclude that Rutin and

Hypersoide are one of the potential drug candidates for

Klebsheilla pneumonia organisms.

The Klebsheilla pneumonia are one of the powerful

pathogens for Multi-Drug Resistance (MDR) in human. The

identified compounds (Quercitrin, Caffeic acid, Chlorogenic

acid, Hyperoside and Rutin) has basically antioxidant activity

whereas these results clearly show that the compounds also

have anti-bacterial properties based on Insilico docking

studies.

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