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Amino Acids

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Design of therapeutically improved analogue of the antimicrobial peptide, indolicidin, using a glycosylation strategy

  • Rohini Dwivedi
  • Priyanka Aggarwal
  • Neel S. Bhavesh
  • Kanwal J. KaurEmail author
Original Article
  • 162 Downloads

Abstract

Indolicidin is a member of cathelicidin family which displays broad spectrum antimicrobial activity. Severe toxicity and aggregation propensity associated with indolicidin pose a huge limitation to its probable therapeutic application. We are reporting the use of glycosylation strategy to design an analogue of indolicidin and subsequently explore structural and functional effects of sugar on it. Our study led to the design of a potent antibacterial glycosylated peptide, [βGlc-T9,K7]indolicidin, which showed decreased toxicity against erythrocytes and macrophage cells and thus a higher therapeutic selectivity. The incorporation of sugar also increased the solubility of the peptide. The mode of bacterial killing, functional stability, LPS binding, and cytokine inhibitory potential of the peptide, however, seemed unaffected upon glycosylation. Absence of significant changes in structure upon glycosylation accounts for the possibly retained functions and mode of action of the peptide. Our report thus presents the designing of an indolicidin analogue with improved therapeutic potential by substituting aromatic amino acid with glycosylated amino acid as a promising strategy for the first time.

Keywords

Antimicrobial peptides Indolicidin Glycosylation Toxicity Therapeutic selectivity Aggregation 

Abbreviations

CARA

Computer-aided resonance assignment

DCC

N,N′-Dicyclohexyl-carbodiimide

DIPEA

Di-isopropyl ethylamine

DMEM

Dulbecco’s modified Eagle’s medium

DMF

Dimethyl formamide

DMSO

Dimethyl sulfoxide

DPC

Dodecylphosphorylcholine

Fmoc

9-Fluorenylmethyloxycarbonyl

HBTU

(2-(1H-benzotraiazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate

HEPES

4-(2-Hydroxyethyl)-1-piperazine ethanesulfonic acid

HOBt

1-Hydroxybenzotriazole

HPLC

High-performance liquid chromatography

IL6

Interleukin 6

LB

Lysogeny broth (Luria–Bertani broth)

LBP

Lipopolysaccharide-binding protein

LPS

Lipopolysaccharide

MALDI-TOF

Matrix-assisted laser desorption and ionization

MHC

Minimum haemolytic concentration

NMP

N-Methyl-2-pyrrolidone

RMSD

Root-mean-square deviation

SEM

Scanning electron microscopy

TOCSY

Total correlation spectroscopy

TNF-α

Tumor necrosis factor alpha

TFE

Tri-fluoroethanol

TFA

Trifluoroacetic acid

Notes

Acknowledgements

We thank Department of Science and Technology (Project no.: SR/S1/OC-63/2012), India for financial support. We thank Department of Biotechnology (DBT), Government of India and ICGEB New Delhi for the NMR facility. We thank Ms. Shanta Sen for help with HRMS data. We also thank Ms. Rekha Rani for help with SEM studies and Dr. Sharad Vashisht for discussions regarding structural studies of glycopeptide.

Author contributions

KK conceived the idea of present work. KK and RD designed the experiments. RD performed the experiments. KK and RD analyzed the data and wrote the manuscript. NSB and PA performed the NMR studies.

Compliance with ethical standards

Conflict of interest

Authors declare that they have no conflicts of interest.

Research involving human participants and/or animals

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

726_2019_2779_MOESM1_ESM.docx (603 kb)
NMR chemical shift values of peptides [T9,K7]indolicidin and [βGlc-T9,K7]indolicidin; PDB and BMRB accession codes of peptides; HPLC profiles of purified indolicidin, [T9,K7]indolicidin and [βGlc-T9,K7]indolicidin; HRMS spectra of indolicidin, [T9,K7]indolicidin and [βGlc-T9,K7]indolicidin; Anti-biofilm activity of peptides (DOCX 603 kb)

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Authors and Affiliations

  1. 1.National Institute of ImmunologyNew DelhiIndia
  2. 2.International Centre for Genetic Engineering and BiotechnologyNew DelhiIndia

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