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LF f(20-31) modified [id=LFH0074]

Producer Organism : Native Protein : Production Method :
Human Lactoferrin (LF) Synthetic
Activity : Antibacterial, antifungal
Target Organisms :

Gram-positive: Staphylococcus aureus MRS 3525 (MMC99=50 g/ml).

Gram-negative: Escherichia coli O6K5 (MMC99=25 g/ml).

Yeast: Candida albicans ATCC 64549 (MMC99=25 g/ml).

Description :
Production method: Synthetic.

Third and seventh residues replaced with alanine, fourth residue replaced with leucine, fifth, sixth and ninth residues replaced with lysine.
Increasing the number of positively charged amino acids by substitution at position 24 (Gln) increased the microbicidal activity (LFH0034). Combining this replacement with the substitution of Asn at position 26 by Ala gave rise to a peptide with significantly increased microbicidal activities for all three microorganisms (LFH0071). Moving Ala substitution from N26 to Q22 (LFH0072) affected the antimicrobial activity negatively compared with that of the former peptide sequence (LFH0071). The two separate Ala substitutions that resulted in significantly enhanced microbicidal activity were introduced into the same peptide (LFH0073). However, the activity was not different from that observed by the Ala substitution at N26 (LFH0036). In addition to the two Ala substitutions, introducing Lys at Gln24 and changing the Arg residues at positions 25 and 28 to Lys also gave rise to a peptide that was significantly better than the natural one against E. coli and S. aureus (LFH0074).
Length : 12 Mass (Da): 1 422.94 Common Amino Acids : K
Isolectric Point : 11.25 Net Charge : 5 Absent Amino Acids : DEGHINPQSTWY
Basic Residues : 5 Acidic Residues : 0 Hydrophobic Residues : 5
Polar Residues : 1 Boman Index : -17.93 Hydropathy Index : -0.108
Aliphatic Index : 73.33 Instability Index : 0 Extinction Coefficient :
Absorbance 280nm : 0

Wheel representation

Hydrophobicity plot

Red solid plot : values according to the hydrophobicity scale of Kyte and Doolittle (reference paper).
Yellow dashed plot : Experimentally determined hydrophobicity scale for proteins at membrane interfaces(reference paper).
Green dotted-dashed plot : prediction of transmembrane helices (reference paper). In this scale (unlike the others), more negative values reflect greater hydrophobicity.

Citation: 1

Structure-microbicidal activity relationship of synthetic fragments derived from the antibacterial alpha-helix of human lactoferrin

Cited Entries: LFH0011, LFH0012, LFH0013, LFH0014, LFH0015, LFH0021, LFH0027, LFH0029, LFH0030, LFH0031, LFH0032, LFH0033, LFH0034, LFH0035, LFH0036, LFH0037, LFH0038, LFH0039, LFH0040, LFH0041, LFH0045, LFH0055, LFH0066, LFH0067, LFH0068, LFH0069, LFH0070, LFH0071, LFH0072, LFH0073, LFH0074, LFH0075, LFH0076, LFH0077, LFH0078

Authors:Haversen, L., Kondori, N., Baltzer, L., Hanson, L.A., Dolphin, G.T., Duner, K., Mattsby-Baltzer, I.
Journal: Antimicrobial Agents and Chemotherapy. 2010, 54(1).
Abstract: There is a need for new microbicidal agents with therapeutic potential due to antibiotic resistance in bacteria and fungi. In this study, the structure-microbicidal activity relationship of amino acid residues 14 to 31 (sequence 14-31) from the N-terminal end, corresponding to the antibacterial {alpha}-helix of human lactoferrin (LF), was investigated by downsizing, alanine scanning, and substitution of amino acids. Microbicidal analysis (99% killing) was performed by a microplate assay using Escherichia coli, Staphylococcus aureus, and Candida albicans as test organisms. Starting from the N-terminal end, downsizing of peptide sequence 14-31 showed that the peptide sequence 19-31 (KCFQWQRNMRKVR, HL9) was the optimal length for antimicrobial activity. Furthermore, HL9 bound to lipid A/lipopolysaccharide, as shown by neutralizing endotoxic activity in a Limulus assay. Alanine scanning of peptide sequence 20-31 showed that Cys20, Trp23, Arg28, Lys29, or Arg31 was important for expressing full killing activity, particularly against C. albicans. Substituting the neutral hydrophilic amino acids Gln24 and Asn26 for Lys and Ala (HLopt2), respectively, enhanced microbicidal activity significantly against all test organisms compared to the amino acids natural counterpart, also, in comparison with HL9, HLopt2 had more than 10-fold-stronger fungicidal activity. Furthermore, HLopt2 was less affected by metallic salts than HL9. The microbicidal activity of HLopt2 was slightly reduced only at pH 7.0, as tested in the pH range of 4.5 to 7.5. The results showed that the microbicidal activity of synthetic peptide sequences, based on the antimicrobial {alpha}-helix region of LF, can be significantly enhanced by optimizing the length and substitution of neutral amino acids at specific positions, thus suggesting a sequence lead with therapeutic potential.

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