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Role of Aromatic Amino Acids in Lipopolysaccharide and Membrane Interactions of Antimicrobial Peptides for use in Plant Disease Control
Bose Inst, Dept Biophys, P-1-12 CIT Scheme 7 M, Kolkata 700054, India. (Anirban Bhunia`s Group)
Bose Inst, Dept Biophys, P-1-12 CIT Scheme 7 M, Kolkata 700054, India. (Anirban Bhunia`s Group)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy. (Martin Malmsten`s group)
Bose Inst, Dept Biophys, P-1-12 CIT Scheme 7 M, Kolkata 700054, India. (Anirban Bhunia)
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2016 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 291, no 25, 13301-13317 p.Article in journal, Meeting abstract (Refereed) Published
Abstract [en]

KYE28(KYEITTIHNLFRKLTHRLFRRNFGYTLR), the representative sequence  of helix D of heparin co-factor II, was demonstrated to be potent against agronomically important Gram-negative plant pathogens X. vesicatoria and X. oryzae,capable of inhibiting disease symptoms in detached tomato leaves. NMR studies in presence of lipopolysaccharide provided structural insights into the mechanisms underlying this, notably in relation to outer membrane permeabilisation. The three-dimensional solution structure of KYE28 in LPS is characterised by a N-ter helical segment, an intermediate loop and an extended C-ter. The two termini are in close proximity to each other via aromatic packing interactions, while the positively charged residues formed an exterior polar shell. To further demonstrate the importance of the aromatic residues for this, a mutant peptide KYE28A, with Ala substitutions at F11, F19, F23 and Y25 showed attenuated antimicrobial activity at high salt concentrations, as well as lower membrane disruption and LPS binding abilities compared to KYE28. In contrast to KYE28, KYE28A adopted an opened out helical structure in LPS with extended N- and C-ter and a small break in between the helical segments. Aromatic packing interactions were completely lost, although hydrophobic interaction between the side chains of hydrophobic residues were still partly retained, imparting an amphipathic character and explaining its residual antimicrobial activity and LPS binding as observed from ellipsometry and ITC. We thus present important structural aspects of KYE28, constituting an aromatic zipper, of potential importance, for the development of novel plant protection agents and therapeutic agents.

Place, publisher, year, edition, pages
2016. Vol. 291, no 25, 13301-13317 p.
Keyword [en]
LPS, Antimicrobial
National Category
Medical and Health Sciences
Research subject
Pharmaceutical Physical Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-282778DOI: 10.1074/jbc.M116.719575ISI: 000379770500033PubMedID: 27137928OAI: oai:DiVA.org:uu-282778DiVA: diva2:917524
Available from: 2016-04-06 Created: 2016-04-06 Last updated: 2017-11-30Bibliographically approved
In thesis
1. Amphiphilic Peptide Interactions with Complex Biological Membranes: Effect of peptide properties on antimicrobial and anti-inflammatory effects
Open this publication in new window or tab >>Amphiphilic Peptide Interactions with Complex Biological Membranes: Effect of peptide properties on antimicrobial and anti-inflammatory effects
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

With increasing problem of resistance development in bacteria against conventional antibiotics, as well as problems associated with diseases either triggered or enhanced by infection, there is an urgent need to identify new types of effective therapeutics for the treatment of infectious diseases and its consequences. Antimicrobial and anti-inflammatory peptides have attracted considerable interest as potential new antibiotics in this context. While antimicrobial function of such peptides is being increasingly understood demonstrated to be due to bacterial membrane disruption, the mechanisms of their anti-inflammatory function are poorly understood. Since bacterial membrane component lipopolysaccharide triggers inflammation, this thesis aims at clarifying importance of lipopolysaccharide (LPS)-peptide interactions while investigating possible modes of action of peptides exhibiting anti-inflammatory effect. Furthermore, effect of poly(ethylene)glycol (PEG)-conjugation was investigated to increase performance of such peptides.

Results presented in this thesis demonstrate that peptide-induced LPS- and lipid A binding/scavenging is necessary but not sufficient criterium for anti-inflammatory effects of peptides. Furthermore, preferential binding to LPS over lipid membrane, as well as higher binding affinity to the lipid A moiety within LPS, are seen for these peptides. In addition, results demonstrate that apart from direct LPS scavenging, membrane-localized peptide-induced LPS scavenging seem to contribute partially to anti-inflammatory effect. Furthermore, fragmentation and densification of LPS aggregates, in turn dependent on the peptide secondary structure on LPS binding, as well as aromatic packing interactions, correlate to the anti-inflammatory effect, thus promoting peptide-induced packing transition in LPS aggregates as key for anti-inflammatory functionality. Thus, peptide-induced LPS aggregate disruption together with reduction of the negative charge of LPS suggests the importance of phagocytosis as an alternative to the inflammatory pathway, which needs to be further investigated. Furthermore, PEG conjugation of peptide results in strongly reduced toxicity at a cost of reduced antimicrobial activity but markedly retained anti-inflammatory effect.

Taken together, the results obtained in this work have demonstrated several key issues which need to be taken into consideration in the development of effective and selective anti-inflammatory peptide therapeutics for the treatment of severe Gram-negative bacterial infections.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 64 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 216
Keyword
LPS, Antimicrobial, Peptide, Inflammation, Infections, Liposome, Binding, PEG
National Category
Medical and Health Sciences
Research subject
Pharmaceutical Physical Chemistry
Identifiers
urn:nbn:se:uu:diva-282781 (URN)978-91-554-9559-6 (ISBN)
Public defence
2016-06-03, B41, BMC, Husargatan 3, Uppsala, Uppsala, 09:15 (English)
Opponent
Supervisors
Funder
Swedish Research Council
Available from: 2016-05-13 Created: 2016-04-06 Last updated: 2016-06-01

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Singh, ShaliniMalmsten, Martin

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