Prediction Of Conserved Hiv Inhibiting Peptide

Published Date: 02 Nov 2017

Abstract

Antipeptide or inhibiting peptide aim to preventing virus/coreceptor interaction. HIV inhibting peptide dataset collected from HIPdb database was used in this study. There are 110 HIV inhibiting peptide is present in HIPdb database. Multiple Sequence Alignment (MSA) of all total 110 antipeptide has been performed and got some best conserved anti-peptides. Next, prediction of antigenicity method was used for finding the maximum antigenicity out of the 14Conserved inhibiting peptides. All peptides were screened for hydrophobicity ,as low hydrophobicity induce humoral mediated immunity. In this study, DRRPASCGTC is showing minimum hydrophobicity with 20% and also having high antigenicity with 0.7480. This study will help scientists to promote research for better understanding of HIV treatment in forms of drug and vaccine development.

Keywords: HIV, HIPdb, Multiple Sequence Alignment, Antigenicity.

1.Introduction

Acquired immunodeficiency syndrome (AIDS) is caused by Human immunodeficiency virus (HIV) in which progressive failure of the human immune system cause life- threating opportunistic infection or cancer. HIV -1 virus multiplies only inside the human body. HIV-1 entry into cells involves formation of a complex between gp120 of the viral envelope glycoprotein (Env), a host receptor (CD4), and a chemokine coreceptors usually CCR5 or CXCR4 [1]. gp 120 contains the transmembrane protein gp41 and derived from polyprotein gp160. Polyprotein gp 160 is encoded by env gene, which is found in all retrovirus [2]. Antiretroviral drugs, peptides have demonstrated potential to inhibit the Human immunodeficiency virus (HIV) [3]. Antipeptide or inhibiting peptide aim to preventing virus/coreceptor interaction by binding either virus envelope proteins or host proteins. The main cause of HIV infection starts with the interaction of exterior envelop of the viral protein gp120 with the chemokine receptors of CD4,the target cell.CD4 not only contributes in the viral attachment but also triggers some conformational changes in the HIV envelop that helps in the recognition of various chemokine receptors and leads to the membrane fusion. These conformational changes leads to the increase in the sensitivity of gp120 loops , release number of gp120 proteins and formation of chemokine receptor site along with its exposure of to gp120 molecules. It also leads to the formation of epitopes of neutralizing antibodies which blocks chemokine receptor binding [4].

Materials and Methods

2.1. Dataset for HIV antipeptide

HIV antipeptide dataset collected from HIPdb [3] database is freely available HIV antipeptide database. It is a manually curated database of experimentally validated HIV inhibitory peptides targeting various steps or proteins involved in the life cycle of HIV like fusion, integration, reverse transcription etc. It is newly introduced database for HIV antipetides, was used for collection of viral entry antipeptides. There are total 110 large dataset of antipeptide having different source and cell lines [Table1] in this database. Out of 110 peptides, 47, 41, 5, 5, 4, 2,2,1,1 inhibiting peptides were taken from GB virus, alpha1-anti trypsin , Apelin,gp120,gp41, synthetic, RhoA, Tat, Dynein sources respectively.

Table 1: HIV inhibiting peptide dataset in HIPdb database

S.No.

ID

SEQUENCE

LENGTH

SOURCE

CELL LINE

HIP1153

FVFLM

5

alpha1-antitrypsin

NA

HIP1159

EFVFLM

6

alpha1-antitrypsin

NA

HIP1158

PFVFLE

6

alpha1-antitrypsin

NA

HIP1160

PEVFLM

6

alpha1-antitrypsin

NA

HIP1157

PFVYLI

6

alpha1-antitrypsin

NA

HIP1154

PFVFLM

6

alpha1-antitrypsin

NA

HIP1161

PFVFLR

6

alpha1-antitrypsin

NA

HIP1155

KPFVFLM

7

alpha1-antitrypsin

NA

HIP1156

NKPFVFLM

8

alpha1-antitrypsin

NA

HIP944

GRKKRRQRRR

10

Tat

P4-R5 MAG

HIP177

PRLSHKGPMPF

11

Apelin

NP-2/CD4

HIP189

RPRLSHKGPMPF

12

Apelin

NP-2/CD4

HIP260

QRPRLSHKGPMPF

13

Apelin

NP-2/CD4

HIP951

PKSSWSDHEASSGV

14

RhoA

TZM-bl

HIP405

KFRRQRPRLSHKGPMPF

17

Apelin

NP-2/CD4

HIP950

TDVILMCFSIDSPDSLENI

19

RhoA

TZM-bl

HIP557

AEAIPMSIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP661

LEAIPMSIPPEVKFNKPAVF

20

alpha1-antitrypsin

P4-CCR5

HIP662

LEAIPMSIPPEVKFNKPFAF

20

alpha1-antitrypsin

P4-CCR5

HIP630

LAAIPMSIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP663

LEAIPMSIPPEVKFNKPFVA

20

alpha1-antitrypsin

P4-CCR5

HIP632

LEAAPMSIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP628

KVINPEPIVEPFMSKPFALF

20

alpha1-antitrypsin

P4-CCR5

HIP638

LEAIPCSIPPCVAFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP635

LEAIPCSIPPCFAFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP639

LEAIPCSIPPCVFFGKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP637

LEAIPCSIPPCFLFGKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP656

LEAIPMSIPPEVFFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP642

LEAIPCSIPPCVGFGKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP643

LEAIPCSIPPCVLFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP640

LEAIPCSIPPCVFFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP646

LEAIPMCIPPECAFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP652

LEAIPMSIPPEFLFGKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP651

LEAIPMSIPPEAKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP659

LEAIPMSIPPEVKFNAPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP650

LEAIPMSIPPAVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP660

LEAIPMSIPPEVKFNKAFVF

20

alpha1-antitrypsin

P4-CCR5

HIP634

LEAIPASIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP664

LEAIPMSIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP648

LEAIPMSIAPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP658

LEAIPMSIPPEVKFAKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP647

LEAIPMSAPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP633

LEAIAMSIPPEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP654

LEAIPMSIPPEVAFAKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP657

LEAIPMSIPPEVKANKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP655

LEAIPMSIPPEVAFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP649

LEAIPMSIPAEVKFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP636

LEAIPCSIPPCFAFNKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP645

LEAIPCSIPPEFLFGKPFVF

20

alpha1-antitrypsin

P4-CCR5

HIP947

GCKKYRRFRWKFKGKFWFWG

20

Synthetic

TZM-bl

HIP948

GKKYRRFRWKFKFGKWFWFG

20

Synthetic

TZM-bl

HIP686

PTGERVWDRGNVTLLCDCPN

20

GB virus C gE2

TZM-bl

HIP694

RIPTGERVWDRGNVTLLCDC

20

GB virus C gE2

TZM-bl

HIP728

WDRGNVTLLCDCPNGPWVWV

20

GB virus C gE2

TZM-bl

HIP600

GPWVWVPAFCQAVGWGDPIT

20

GB virus C gE2

TZM-bl

HIP712

TLLCDCPNGPWVWVPAFCQA

20

GB virus C gE2

TZM-bl

HIP631

LCDCPNGPWVWVPAFCQAVG

20

GB virus C gE2

TZM-bl

HIP569

DCPNGPWVWVPAFCQAVGWG

20

GB virus C gE2

TZM-bl

HIP684

PNGPWVWVPAFCQAVGWGDP

20

GB virus C gE2

TZM-bl

HIP730

WVWVPAFCQAVGWGDPITHW

20

GB virus C gE2

TZM-bl

HIP592

GAPASVLGSRPFDYGLKWQS

20

GB virus C gE2

TZM-bl

HIP724

VSVTCVWGSVSWFASTGGRD

20

GB virus C gE2

TZM-bl

HIP702

SWFASTGGRDSKIDVWSLVP

20

GB virus C gE2

TZM-bl

HIP699

SKIDVWSLVPVGSASCTIAA

20

GB virus C gE2

TZM-bl

HIP717

VGSASCTIAALGSSDRDTVV

20

GB virus C gE2

TZM-bl

HIP665

LGSSDRDTVVELSEWGVPCV

20

GB virus C gE2

TZM-bl

HIP580

ELSEWGVPCVTCILDRRPAS

20

GB virus C gE2

TZM-bl

HIP704

TCILDRRPASCGTCVRDCWP

20

GB virus C gE2

TZM-bl

HIP563

CGTCVRDCWPETGSVRFPFH

20

GB virus C gE2

TZM-bl

HIP585

ETGSVRFPFHRCGTGPRLTK

20

GB virus C gE2

TZM-bl

HIP691

RCGTGPRLTKDLEAVPFVNR

20

GB virus C gE2

TZM-bl

HIP681

PFDYGLKWQSCSCRANGSRI

20

GB virus C gE2

TZM-bl

HIP575

DLEAVPFVNRTTPFTIRGPL

20

GB virus C gE2

TZM-bl

HIP716

TTPFTIRGPLGNQGRGNPVR

20

GB virus C gE2

TZM-bl

HIP599

GNQGRGNPVRSPLGFGSYTM

20

GB virus C gE2

TZM-bl

HIP701

SPLGFGSYTMTKIRDSLHLV

20

GB virus C gE2

TZM-bl

HIP711

TKIRDSLHLVKCPTPAIEPP

20

GB virus C gE2

TZM-bl

HIP614

KCPTPAIEPPTGTFGFFPGV

20

GB virus C gE2

TZM-bl

HIP708

TGTFGFFPGVPPINNCMPLG

20

GB virus C gE2

TZM-bl

HIP685

PPINNCMPLGTEVSEALGGA

20

GB virus C gE2

TZM-bl

HIP707

TEVSEALGGAGLTGGFYEPL

20

GB virus C gE2

TZM-bl

HIP598

GLTGGFYEPLVRRCSELMGR

20

GB virus C gE2

TZM-bl

HIP566

CSCRANGSRIPTGERVWDRG

20

GB virus C gE2

TZM-bl

HIP723

VRRCSELMGRRNPVCPGYAW

20

GB virus C gE2

TZM-bl

HIP696

RNPVCPGYAWLSSGRPDGFI

20

GB virus C gE2

TZM-bl

HIP669

LSSGRPDGFIHVQGHLQEVD

20

GB virus C gE2

TZM-bl

HIP565

CRANGSRIPTGERVWDRGNV

20

GB virus C gE2

TZM-bl

HIP560

ANGSRIPTGERVWDRGNVTL

20

GB virus C gE2

TZM-bl

HIP603

GSRIPTGERVWDRGNVTLLC

20

GB virus C gE2

TZM-bl

HIP593

GERVWDRGNVTLLCDCPNGP

20

GB virus C gE2

TZM-bl

HIP698

RVWDRGNVTLLCDCPNGPWV

20

GB virus C gE2

TZM-bl

HIP676

NVTLLCDCPNGPWVWVPAFC

20

GB virus C gE2

TZM-bl

HIP729

WVPAFCQAVGWGDPITHWSH

20

GB virus C gE2

TZM-bl

HIP677

PAFCQAVGWGDPITHWSHGQ

20

GB virus C gE2

TZM-bl

HIP588

FCQAVGWGDPITHWSHGQNQ

20

GB virus C gE2

TZM-bl

HIP687

QAVGWGDPITHWSHGQNQWP

20

GB virus C gE2

TZM-bl

HIP608

HWSHGQNQWPLSCPQYVYGS

20

GB virus C gE2

TZM-bl

HIP668

LSCPQYVYGSVSVTCVWGSV

20

GB virus C gE2

TZM-bl

HIP693

RGNVTLLCDCPNGPWVWVPA

20

GB virus C gE2

TZM-bl

HIP946

PKDGPSPGGTLMDLSERQEVS

SVRSLSST

29

Dynein

HeLa

HIP824

LVQPRGPRSGPGPWQGGRRK

FRRQRPRLSHKGPMPF

36

Apelin

NP-2/CD4

HIP963

CTRPNNNTRKSIRIQRGPGRAF

VTIGKIGNMRQAHC

36

gp120

JY

HIP958

YTSLIHSLIEESQNQQEKNEQEL

LELDKWASLWNWF

36

gp41

PM-1

HIP959

YTSLIHSLIEESQNQQEKNEQEL

LELDKWASLANAA

36

gp41

PM-1

HIP965

EINCTRPNNNTRKSIHIGPGRAF

YTTGEIIGDIRQAHCNIS

41

gp120

JY

HIP962

EINCTRPNNNTRKSIRIQRGPG

RAFVTIGKIGNMRQAHCNIS

42

gp120

JY

HIP964

ESVKITCARPYQNTRQRTPIGL

GQSLYTTRSRSIIGQAHCNIS

43

gp120

JY

HIP966

ESVVINCTRPNNNTRRRLSIGP

GRAFYARRNIIGDIRQAHCNIS

44

gp120

JY

HIP1016

WMEWDREINNYTSLIHSLIEES

QNQQEKNEQELLELDKWASLWNWFRS

48

gp41

PM-1

HIP953

WMEWDREINNYTSLIHSLIEESQ

NQQEKNEQELLELDKWASLWNWFRS

48

gp41

PM-1

Collection of the HIV inhibiting peptide data set from HIPdb Database

Performed Multiple sequence alignment of all 110 HIV inhibiting peptide

Prediction of antigenicity of conserved peptides and hydrophobicity

Selection of maximum antigenicity with minimum hydrophobicity of HIV inhibiting peptide

Fig. 1. Overall block diagram of the methodology used

Multiple sequence alignment (MSA):

MSA were perfomed for all 110 inhibiting peptides using clustalW [5] and validate its result with Tcoffee [6] and Muscle [7]. Conserved patches of antipeptides for different cell lines and sources were collected from the clustalW result.

Prediction of Antigenicity of Conserved peptides and analysis

Low hydrophobic sequence evokes the production of peptide antibodies. The Antigenicity analysis assist in the selection of low hydrophobic sequences from the one with the highest calculated antigenic potential. All the antipeptides were screened for their antigenicity using Vaxijen [8]. The threshold value for antigenicity is 0.4 in vaxijen. Based on their conserved nature and antigenicity, few lists of peptides are proposed for their wide spectrum activity. The antigenicity is the ability of a compound to bind with antibodies and with cells of immune system.

2.4 Prediction of Antigenicity of Conserved peptides and analysis

Peptide property calculator [9] was used to calculate the percentage of hydrophobicity of all antipeptides. By using this tool, we got chemical formulae, molecular weight, hydrophobicity, hydrophilic, isoelectric point for analysis of inhibiting peptides. We analyzed our whole 11 inhibiting peptide with derived ID from HIPDB database inhibiting from Peptide Property Calculator.

2.5 Prediction of Antiviral and antimicrobial activity of 11 peptides:

Conserved Antimicrobial Peptide (CAMP) [10] is used for prediction of antimicrobial activity of our 11 peptides using the Support Vector Machine (SVM) [ 11] and Random Forest (RF) [12] algorithm. The category of AMPs has been analysed from class AMP [13].

Results:

The results of MSA of 14 antipeptides were visualized in Jalview. Fig2 is showing the decamers antipeptides in jalview. Then we peredicted antigenicity of all 14 inhibiting peptides through Vaxijen. All the 14 HIV antipeptides antigenicity is shown in table 2. The final step is to predict the best HIV antipeptide among these 14 as shown below (table3). In our result DRRPASCGTC, EESQNQQEKN, KYRRFRWKFK, SDRDTVVELS and KIRPSLHLVKC are showing hydrophobicity below 60%. These antipeptides have potential to inhibits HIV virus entry and they are able to induce humoral mediated immunity . In this study, DRRPASCGTC is showing minimum hydrophobicity with 20% and also having high antigenicity with 0.7480.

Fig 2: Jalview image of decamers inhibiting peptides.

Table 2: All the 14 HIV antipeptides antigenicity

S.No.

ID

Sequence

SOURCE

CELL LINE

HIP947

GCKKYRRFRWKFKGKFWFWG

Synthetic

TZM-bl

HIP948

GKKYRRFRWKFKFGKWFWFG

Synthetic

TZM-bl

HIP686

PTGERVWDRGNVTLLCDCPN

GB virus C gE2

TZM-bl

HIP694

RIPTGERVWDRGNVTLLCDC

GB virus C gE2

TZM-bl

HIP665

LGSSDRDTVVELSEWGVPCV

GB virus C gE2

TZM-bl

HIP704

TCILDRRPASCGTCVRDCWP

GB virus C gE2

TZM-bl

HIP599

GNQGRGNPVRSPLGFGSYTM

GB virus C gE2

TZM-bl

HIP711

TKIRDSLHLVKCPTPAIEPP

GB virus C gE2

TZM-bl

HIP824

LVQPRGPRSGPGPWQGGRRKFRRQRPRLSHKGPMPF

Apelin

NP 2/CD4

HIP958

YTSLIHSLIEESQNQQEKNEQELLELDKWASLWNWF

gp41

PM-1

HIP965

EINCTRPNNNTRKSIHIGPGRAFYTTGEIIGDIRQAHCNIS

gp120

JY

HIP964

ESVKITCARPYQNTRQRTPIGLGQSLYTTRSRSIIGQAHCNIS

gp120

JY

HIP966

ESVVINCTRPNNNTRRRLSIGPGRAFYARRNIIGDIRQAHCNIS

gp120

JY

HIP1016

WMEWDREINNYTSLIHSLIEESQNQQEKNEQELLELDKWASLWNWFRS

gp41

PM-1

Table 3: The best HIV antipeptide

Conserve Hiv Anti- Peptide Patches

Antigen (Threshold=0.4)

H*

HIPdb antipeptide

Antigen (Threshold=0.4)

H*

Source

Cell- Line

EESQNQQEKN (2times)

Overall Antigen Prediction = 0.5418 ( Probable ANTIGEN ).

0%

YTSLIHSLIEESQNQQEKNEQELLELDKWASLWNWF

-0.0022

36

gp41

PM-1

PTGERVWDRG(2 times)

Overall Antigen Prediction = 0.2602 ( Probable NON-ANTIGEN ).

30%

PTGERVWDRGNVTLLCDCPN

0.3447

35

GB Virus C gE2

TZM-bl

KYRRFRWKFK (2 times)

Overall Antigen Prediction = 1.6162 ( Probable ANTIGEN ).

60%

GCKKYRRFRWKFKGKFWFWG

1.2550

35

Synthetic

TZM-bl

PNNNTRKSIH

Overall Antigen Prediction = 0.1556 ( Probable NON-ANTIGEN ).

20%

EINCTRPNNNTRKSIRIQRGPGRAFVTIGKIGNMRQAHCNIS

0.6019

31

gp120

JY

PNNNTRRRLS

Overall Antigen Prediction = -0.3115 ( Probable NON-ANTIGEN ).

20%

ESVKITCARPYQNTRQRTPIGLGQSLYTTRSRSIIGQAHCNIS

0.613

34

gp120

JY

PYQNTRQRTP

Overall Antigen Prediction = 0.1124 ( Probable NON-ANTIGEN ).

20%

ESVKITCARPYQNTRQRTPIGLGQSLYTTRSRSIIGQAHCNIS

0.7910

28

gp120

JY

PWQGGRRKFR

Overall Antigen Prediction = 0.1293 ( Probable NON-ANTIGEN ).

30%

LVQPRGPRSGPGPWQGGRRKFRRQRPRLSHKGPMPF

0.3389

39

Apelin

NP-2/CD4

RGNPVRSPLG

Overall Antigen Prediction = -0.2856 ( Probable NON-ANTIGEN ).

40%

GNQGRGNPVRSPLGFGSYTM

0.6627

30

GB Virus C gE2

TZM-bl

SDRDTVVELS

Overall Antigen Prediction = 0.5728 ( Probable ANTIGEN ).

30%

LGSSDRDTVVELSEWGVPCV

0.7667

40

GB Virus C gE2

TZM-bl

KIRDSLHLVKC

Overall Antigen Prediction = 0.5728 ( Probable ANTIGEN ).

45%

TKIRDSLHLVKCPTPAIEPP

0.1542

50

GB Virus C gE2

TZM-bl

DRRPASCGTC

Overall Antigen Prediction = 0.7480 ( Probable ANTIGEN ).

20%

TCILDRRPASCGTCVRDCWP

1.3830

35

GB Virus C gE2

TZM-bl

Note: H* indicates hydrophobocity.

Table 4: Hydrophobicity of 11 conserved anti-peptides:

Peptide Sequence

H** (%)

H*(%)

Others (%)

Chemical Formula

MW(Da)

pI

Piechart

KIRDSLHLVKC

45

36

18

C57H102N18O15S1

1311.60

9.67

KYRRFRWKFK

60

30

10

C74H111N23O12

1514.83

12.27

PNNNTRKSIH

30

20

50

C48H81N19O16

1180.28

11.66

PNNNTRRRLS

30

20

50

C48H86N22O16

1227.34

12.80

PTGERVWDRG

40

30

30

C50H77N17O16

1172.26

6.51

PWQGGRRKFR

40

30

30

C58H90N22O12

1287.48

12.81

RGNPVRSPLG

20

40

40

C44H77N17O13

1052.19

12.50

SDRDTVVELS

40

30

30

C45H77N13O20

1120.17

3.88

EESQNQQEKN

40

0

60

C47H76N16O23

1233.21

3.98

DRRPASCGTC

30

20

50

C39H68N16O15S2

1065.19

8.23

PYQNTRQRTP

20

20

60

C53H85N19O17

1260.36

11.15

Note: H* indicates hydrophobicity ; pI indicates isoelectric point; H** indicates hydrophilicity; MW indicates molecular weight.

Piechart : Red indicates +vely charged hydrophilic residues; Blue indicates -vely charged hydrophilic residues;

Green indicates hydrophobic residues; Grey indicates others.

Table 5:Conserved antimicrobial peptides

S. No.

Antipeptides

Class(SVM)

Probabilty(SVM)

Class(RF)

Probabilty(RF)

1

EESQNQQEKN

AMP

0.696

AMP

0.55

2

PTGERVWDRG

Non-AMP

0.706

AMP

0.706

3

KYRRFRWKFK

AMP

1.000

AMP

0.892

4

PNNNTRKSIH

AMP

0.913

Non-AMP

0.586

5

PNNNTRRRLS

AMP

0.589

Non-AMP

0.598

6

PYQNTRQRTP

Non-AMP

0.993

AMP

0.528

7

PWQGGRRKFR

AMP

0.981

AMP

0.81

8

RGNPVRSPLG

AMP

0.625

AMP

0.632

9

SDRDTVVELS

Non-AMP

0.959

AMP

0.624

10

KIRDSLHLVKC

AMP

0.588

AMP

0.712

11

DRRPASCGTC

AMP

0.678

AMP

0.604

Table 6: AMP analysis of Conserved antipeptides

Peptide Sequence

Prediction method- SVM

Prediction method- RF

Prediction

Probability

Prediction

Probability

KIRDSLHLVKC

Antifungal

0.98007525738161

Antibacterial

0.858

KYRRFRWKFK

Antibacterial

0.997191821246762

Antibacterial

0.564

PNNNTRKSIH

Antifungal

0.716303607753123

Antifungal

0.562

PNNNTRRRLS

Antiviral

0.708622545527062

Antifungal

0.63

PTGERVWDRG

Antiviral

0.757892210674696

Antibacterial

0.498

PWQGGRRKFR

Antiviral

0.95167536018741

Antibacterial

0.654

RGNPVRSPLG

Antiviral

0.953364870196997

Antibacterial

0.772

SDRDTVVELS

Antifungal

0.789076193791002

Antibacterial

0.372

EESQNQQEKN

Antifungal

0.727395582704812

Antifungal

0.586

DRRPASCGTC

Antiviral

0.890473933052895

Antibacterial

0.532

PYQNTRQRTP

Antiviral

0.858966097668196

Antibacterial

0.636

Table 7: Antipeptide and target organism

Antipeptides

Target Organism

sequences

activity

Identity %

E-value

EESQNQQEKN

Simian Immunodeficiency virus

WQEWERKVDFLEENITALLEEAQIQQEKNMYELQK

Antiviral

80

1.3

PTGERVWDRG

E. coli ( ED50 = 30-35 nM ),S.aureus ( ED50 = 90-120 nM )

GLRKKFRKTRKRIQKLGRKIGKTGRKVWKAWREYGQIPYPCRI

Antibacterial

66

24

KYRRFRWKFK

E. coli ATCC 25922 ( MIC = 2 ??M),E. coli ML35 ( MIC =4 ??M),E. coli D21 ( MIC = 4 ??M),S. typhimurium ATCC 14028 ( MIC = 4 ??M),P. aeruginosa ATCC 27853 ( MIC = 1 ??M),S. marcescens ATCC 8100 ( MIC = 2 ??M),S. aureus ATCC 25923 ( MIC = 2 ??M),S. aureus Cowan 1 ( MIC = 2 ??M),S. aureus MRSA ( MIC = 4 ??M),S. epidermidis ATCC 12228 ( MIC = 1 ??M),B. megaterium Bm11 ( MIC = 2 ??M),C. albicans ( MIC = 16 ??M),C. neoformans ( MIC = 4 ??M)

GRFKRFRKKFKKLFKKLS

Antibacterial, Antifungal

60

2.8

PNNNTRKSIH

Unknown

MSRSLKKGPFVFYSLIKKVDQMNSNRFKSVILTWSRSCTIIPIMIGNTIGVYNGKEHIPVLVSDQMIGHKLGEFVQTRNYRGHKKHDKKTKTKR

Antimicrobial

50

155

PNNNTRRRLS

Unknown

SLSVEAKAKIVADFGRDANDTGSSEVQVALLTAQINHLQGHFSEHKKDHHSRRGLLRMVSTRRKLLDYLKRKDVASYVSLIERLGLRR

Antimicrobial

60

119

PYQNTRQRTP

E. coli

DEKPKLILPTPAPPNLPQLVGGGGGNRKDGFGVSVDAHQKVWTSDNGGHSIGVSPGYSQHLPGPYGNSRPDYRIGAGYSYNF

Antibacterial

66

70

PWQGGRRKFR

Unknown

RIRRPIALIWRGGRRLTEWL

Antimicrobial

83

4.8

RGNPVRSPLG

Unknown

RFRPPIRRPPIRPPFRPPFRPPVRPPIRPPFRPPFRPPIGPFP

Antimicrobial

57

53

SDRDTVVELS

Unknown

MKRNQRKQLIGTVVSTKNAKTATVKVTSRFKHPLYHKSVIRHKKYHVHNFGELVANDGDRVQIIETRPLSALKRWRIVKIIERAK

Antimicrobial

42

346

KIRDSLHLVKC

Odorrana grahami (Yunnanfu frog )

GLLSGILGAGKHIVCGLSGPCQSLNRKSSDVEYHLAKC

Antibacterial, Antifungal

80

45

DRRPASCGTC

Unknown

EQKQGQYGEGSLRPSECGQRCSYRCSATSHKKPCMFFCQKCCAKCLCVPPGTFGNKQVCPCYNNWKTQQGGPKCP

Antimicrobial

66

24

Discussion

To explore AMPs as drugs it is essential to understand sequence-specificity relationship of Anti Microbial Peptides (AMPs). Two algorithms has been used for prediction of antibacterial, antifungal and antiviral peptides based on their sequence composistion. The individual inference about each 11 peptides is as follows:

EESQNQQEKN: For a peptide to be antigenic, it must be less hydrophobic and vaxijen score should be more than 0.4. To our surprise, this peptide is predicted to be 0% hydrophobicity and has antigenic score of 0.5418. This peptide has a good probability of immunogenicity because it has molecular weight greater than 500 D, it has moleculer weight of 1233.21. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 0.69 and 0.55 respectively. This peptide has AMP category of Antifungal activity from both algorithms predicted from classAMP, and Random Forest with probability of 0.72 and 0.58 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???WQEWERKVDFLEENITALLEEAQIQQEKNMYELQK??? sequence which has antiviral activity to Simian immunodeficiency Virus with 80% identity & E value of 1.3. Thus this conserved peptide has derived from gp41 sequence which is working as a potent viral entry inhibitor of HIV-1, SIVmac251 and SHIV89.6P [14]. This peptide might work against either CD4/coreceptor or gp120 to inhibit viral entry.

PTGERVWDRG: This peptide is predicted to be 30% hydrophobicity and has antigenic score of 0.2602. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D, it has moleculer weight of 1172.26. As per CAMP analysis, this peptide is predicted to be non-AMP from SVM and AMP from random forest algorithms, with probability of 0.70 and 0.70 respectively. This peptide has AMP category of Antiviral activity from SVM and antibacterial activity from random forest algorithms predicted from classAMP, with probability of 0.75 and 0.49 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???GLRKKFRKTRKRIQKLGRKIGKTGRKBWKAWREYGQIEYPCRI??? sequence which has antibacterial activity to E.coli and S. aureus with 66% identity & E value of 24. This peptide is predicted to be antiviral and antibacterial activity with no immunogenicity. This conserved peptide has been derived from GB Virus C gE2, the peptides P6-2 and P4762 inhibits HIV-1 replication via interaction with HIV-1 particle and avoid the entry of virions [15]. This peptide is sowing antiviral activity with no immunogenicity.

KYRRFRWKFK:

This peptide is predicted to be 30% hydrophobicity and has antigenic score of 1.61. This peptide has a good probability of immunogenicity because it has molecular weight greater than 500 D, it has moleculer weight of 1514.83. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 1 and 0.89 respectively. This peptide has AMP category of Antibacterial activity from both algorithms predicted from classAMP, and Random Forest with probability of 0.99 and 0.56 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???GRFKRFRKKFKKLFKKLS??? sequence which has antibacterial and antifungal activity to E.coli, S.typhimurium, T. aeruginosa, S. marcescens, S. aureus, S. epidermidis, B. megaterium, C. albicans, C. neoformans with 60% identity & E value of 2.8. This conserved peptide is derived from synthetic peptides (SALPs- Synthetic anti-lipopolysaccharides peptides), bind to heparin sulphate moeties on the cell surface and inhibit the entry of HIV-1, HSV-1&2 both, HBV and HCV [16]. These peptides has high antiviral efficiency, no toxicity and adverse effects.This peptide is best as it is showing antibacterial as well as antifungal activity with good score of antigenicity and immunogenicity. This peptide has wide spectrum future prospectives in HIV treatment.

PNNNTRKSIH: This peptide is predicted to be 20% hydrophobicity and has antigenic score of -0.155. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D,, it has moleculer weight of 1180.28. As per CAMP analysis, this peptide is predicted to be AMP from SVM and non-AMP from random forest algorithms, with probability of 0.913 and 0.586 respectively. This peptide has AMP category of Antifungal activity from both algorithms predicted from classAMP, and Random Forest with probability of 0.716 and 0.562 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???MSRSLKKGPFVFYSLIKKVDQMNSNRFKSVILTWSRSCTIIPIMIGNTIGVYNGKEHIPVLVSDQMIGHKLGEFVQPRNYRGHKKHDKKTKTKR??? sequence which has antimicrobial activity to Simian immunodeficiency Virus with 50% identity & E value of 155. Similar to 2 peptide, This peptide is predicted to be antifungal and antimicrobial activity with no immunogenicity. This conserved peptide has been derived from gp120, V3 region of gp120 of T cell line trophic directly interact with CXCR4, a chemokine receptor of CD4+ cells, hence inhibiting T trophic HIV-1 infection [17]. This conserved peptide may be good candidate to inhibit viral entry.

PNNNTRRRLS: This peptide is predicted to be 20% hydrophobicity and has antigenic score of -0.3115. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D,, it has moleculer weight of 1227.34. As per CAMP analysis, this peptide is predicted to be AMP from SVM and non-AMP from random forest algorithms, with probability of 0.589 and 0.598 respectively. This peptide has AMP category of Antiviral activity from SVM and antifungal activity from random forest algorithms predicted from classAMP, with probability of 0.708 and 0.630 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???SLSVEAKAKIVADFGRDANDTGSSEVQVALLTAQINHLQGHFSEHKKDHHSRRGLLRMVSTRRKLLDYLKRKDVASYVSLIERLGLRR??? sequence which has antimicrobial activity with 60% identity & E value of 119. Similar to peptide 4, This peptide is predicted to be antifungal and antimicrobial activity with no immunogenicity. This conserved peptide has been derived from gp120, V3 region of gp120 of T cell line trophic directly interact with CXCR4, a chemokine receptor of CD4+ cells, hence inhibiting T trophic HIV-1 infection .This conserved peptide may be good candidate to inhibit viral entry.

PYQNTRQRTP: This peptide is predicted to be 20% hydrophobicity and has antigenic score of 0.112. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D, it has moleculer weight of 1260.36. As per CAMP analysis, this peptide is predicted to be non-AMP from SVM and AMP from random forest algorithms, with probability of 0.993 and 0.528 respectively. This peptide has AMP category of Antiviral activity from SVM and antibacterial activity from random forest algorithms predicted from classAMP, with probability of 0.858 and 0.636 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???DEKPKLILPTPAPPNLPQLVGGGGGNRKDGFGVSVDAHQKVWTSDNGGHSIGVSPGYSQHLPGPYGNSRPDYRIGAGYSYNF??? sequence which has antibacterial activity to E. coli with 66% identity & E value of 70. This peptide sequence has also the capability to acts as AMP without having the property of inducing immune response. This peptide is predicted to be antiviral and antibacterial activity with no immunogenicity. This conserved peptide has been derived from gp120, V3 region of gp120 of T cell line trophic directly interact with CXCR4, a chemokine receptor of CD4+ cells, hence inhibiting T trophic HIV-1 infection .This conserved peptide may be good candidate to inhibit viral entry.

PWQGGRRKFR: This peptide is predicted to be 30% hydrophobicity and has antigenic score of 0.129. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D, it has moleculer weight of 1287.48. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 0.981 and 0.810 respectively. This peptide has AMP category of Antiviral activity from SVM and antibacterial activity from random forest algorithms predicted from classAMP, with probability of 0.951 and 0.654 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???RIRRPIALIWRGGRRLTEWL??? sequence which has antimicrobial activity with 83% identity & E value of 4.8. This conserved peptide has been derived from Apelin, inhibits the entry of some HIV-1&2 into CD4+ cells APJ receptor [18]. This is another best peptide candidate having antiviral activity with no immunogenic response.

RGNPVRSPLG: This peptide is predicted to be 40% hydrophobicity and has antigenic score of -0.2856. This peptide has no probability of immunogenicity because vaxijen score is less than 0.4, less than threshold, it has molecular weight greater than 500 D, it has moleculer weight of 1052.19. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 0.625 and 0.632 respectively. . This peptide has AMP category of Antiviral activity from SVM and antibacterial activity from random forest algorithms predicted from classAMP, with probability of 0.953 and 0.772 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???RFRPPIRRPPIRPPFRPPFRPPVRPPIRPPFRPPFRPPIGPFP??? sequence which has antimicrobial activity with 57% identity & E value of 53. This conserved peptide has been derived from GB Virus C gE2, the peptides P6-2 and P4762 inhibits HIV-1 replication via interaction with HIV-1 particle and avoid the entry of virions. This peptide is sowing high antiviral activity with no immunogenicity.

SDRDTVVELS: This peptide is predicted to be 30% hydrophobicity and has antigenic score of 0.572. This peptide has probability of immunogenicity because it has molecular weight greater than 500 D, it has moleculer weight of 1120.17. As per CAMP analysis, this peptide is predicted to be non-AMP from SVM and AMP from random forest algorithms, with probability of 0.959 and 0.624 respectively. This peptide has non AMP category of Antifungal activity from SVM and AMP antibacterial activity from random forest algorithms predicted from classAMP, with probability of 0.789 and 0.372 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???MKRNQRKQLIGTVVSTKNAKTATVKVTSRFKHPLYHKSVIRHKKYHVHNFGELVANDGDRVQIIETRPLSALKRWRIVKIIERAK??? sequence which has antimicrobial activity with 42% identity & E value of 346. This conserved peptide has been derived from GB Virus C gE2, the peptides P6-2 and P4762 inhibits HIV-1 replication via interaction with HIV-1 particle and avoid the entry of virions. This peptide is sowing low antiviral activity with immunogenicity.

KIRDSLHLVKC: This peptide is predicted to be 36% hydrophobicity and has antigenic score of 0.572. This peptide has probability of immunogenicity because it has molecular weight greater than 500 D, it has moleculer weight of 1311.60. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 0.588 and 0.712 respectively. This peptide has AMP category of Antifungal activity from SVM and antibacterial activity from Random forest, predicted from classAMP, and Random Forest with probability of 0.980 and 0.858 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???GLLSGILGAGKHIVCGLSGPCQSLNRKSSDVEYHLAKC??? sequence which has antibacterial activity to Odorrana grahami with 80% identity & E value of 45. This conserved peptide has been derived from GB Virus C gE2, the peptides P6-2 and P4762 inhibits HIV-1 replication via interaction with HIV-1 particle and avoid the entry of virions. This peptide is sowing high antiviral activity with immunogenicity.

DRRPASCGTC: This peptide is predicted to be 20% hydrophobicity and has antigenic score of 0.748. This peptide has a good probability of immunogenicity because it has molecular weight greater than 500 D, it has moleculer weight of 1065.19. As per CAMP analysis, this peptide is predicted to be AMP from both algorithms, SVM and Random Forest with probability of 0.678 and 0.604 respectively. This peptide has AMP category of Antiviral activity from SVM and antibacterial activity from Random forest, predicted from classAMP, with probability of 0.890 and 0.532 respectively. After AVBlast analysis of this peptide, it was found that this sequence is a part of ???EQKQGQYGEGSLRPSECGQRCSYRCSATSHKKPCMFFCQKCCAKCLCVPPGTFGNKQVCPCYNNWKTQQGGPKCP??? sequence which has antimicrobial activity to Simian immunodeficiency Virus with 66% identity & E value of 24. This conserved peptide has been derived from GB Virus C gE2, the peptides P6-2 and P4762 inhibits HIV-1 replication via interaction with HIV-1 particle and avoid the entry of virions. This peptide is sowing moderate antiviral activity with high immunogenicity.

In general, peptide sequences between 10 and 20 amino acids in length are recommended for ideal antigen [19] So, our conserved antipeptide having 10 amino acid lengh which can use for inhibitor.In general, most ideal antigenic epitopes are hydrophilic, surface orientated and flexible. Hydrophilic residues are surface exposed so its has better affinity to bind with paratope as compared to hydrophobic residues.

In this study, we also compare hydrophobicity and antigenicity of our conserved antipeptide with antipeptide from HIPdb databases. Low hydrophobicity shows low affinity binding characteristics.

Conclusion

Many efforts are being made for inhibition of HIV virus entry block by insilico, invivo, insitu and invitro approaches. We come on conclusion that DRRPASCGTC is showing minimum hydrophobicity with 20% and high antigenicity with 0.7480.Out of 11 we got 10 having less hydrophicity percentage than derived HIPDB database antipeptide, which proves that our inhibiting peptide will be effective inhibitor peptides than HIPdb antipeptides. These antipeptides have potential to inhibit HIV virus entry and they are able to induce humoral mediated immunity. These are the conserved patches taken from different sources and effective against different cell lines so these are wide spectrum antipeptides inhibiting the primary interaction of gp120-CD4 that is the major culprit for HIV pathogenesis.

Acknowledgements

We are very thankful to the Nthyrs Biotech Labs, 1-8-747/11, Baghlingampally, near RTC Road, Hyderabad-44, where this work was performed. We again acknowledge the Nthrys Biotech Labs for providing infrastructural facilities. Big thanks to all our Nthrys???s scientific staff.

Author Disclosure Statement

No competing financial interests exist.