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1.
Anal Bioanal Chem ; 398(7-8): 3073-9, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20924563

RESUMO

Four novel alanine-based indolicidin peptide derivatives were designed containing one WPW motif and two alanine residues, resulting in peptides of similar sequence. The separation of these peptides with identical physicochemical properties including molar mass, charge, and secondary structure as characterized by circular dichroism spectroscopy is very difficult; and the separation of peptides with differing physicochemical properties has only previously been reported. Capillary electrophoresis parameters such as separation buffer concentration, separation buffer pH, capillary length, and separation voltage were investigated to optimize the analysis. Using optimized conditions of a background electrolyte containing 5 mM formic acid of pH 2.0, total capillary length of 51 cm and a voltage of 10 kV enabled a baseline separation of the four peptides. The relative standard deviation of the peak areas and migration times for method repeatability (n = 3) were found to be lower than 8% and 3%, respectively. In addition, reasoning for the separation of these peptides is proposed based on the acidity of the formic acid buffer and the hydrophobic grouping of the tryptophan residues in the peptide primary sequence.


Assuntos
Alanina/análogos & derivados , Anti-Infecciosos/síntese química , Peptídeos Catiônicos Antimicrobianos/síntese química , Eletroforese Capilar/métodos , Alanina/síntese química , Alanina/química , Alanina/isolamento & purificação , Motivos de Aminoácidos , Anti-Infecciosos/química , Anti-Infecciosos/isolamento & purificação , Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
2.
Org Biomol Chem ; 8(7): 1679-87, 2010 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-20237682

RESUMO

In the current study, indolicidin, a known antimicrobial originally isolated from bovine neutrophils, was modified with respect to hydrophobicity and tryptophan content to maximize bioactivity, and minimize cytotoxicity. Since indolicidin contains five tryptophans (very hydrophobic) of its total 13 amino acids, alanine (mildly hydrophobic) was incrementally substituted in its place to generate five novel derivatives with decreasing hydrophobicity. Antimicrobial testing identified two active derivatives with minimum inhibitory concentrations in the 10(-9) g mL(-1) range against Candida albicans, as well as broad-spectrum activity against various other Gram-positive/negative pathogens in the 10(-3)-10(-6) g mL(-1) range. Cytotoxicity testing yielded minimum hemolytic concentrations of approximately 3 x 10(-3) g mL(-1) for both active derivatives, resulting in hemolytic indices of >1.3 x 10(6) (peptide Delta45) and 3.6 x 10(5) (peptide Delta5) (improvements of >33,000-fold and approximately 10,000-fold, respectively, compared to indolicidin). The potent antimicrobial activity and low cytotoxicity of these derivatives show promise as potential antibiotics.


Assuntos
Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias/efeitos dos fármacos , Candida albicans/efeitos dos fármacos , Triptofano/química , Sequência de Aminoácidos , Interações Hidrofóbicas e Hidrofílicas , Testes de Sensibilidade Microbiana
3.
Bioorg Chem ; 38(3): 98-107, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20153500

RESUMO

The capping efficiency of glycine on cavitand-based synthetic four-helix bundles was investigated. Glycine, a common C-capping amino acid, has always been included as a C-terminal residue in our de novo peptides, although the exact contribution of the glyince cap to the overall stability and structure of the caviteins had not previously been examined. The uncapped proteins were found to be less helical according to their CD spectra. In addition, the H/D exchange experiments suggested that the uncapped caviteins were more conformationally flexible. Capped and uncapped caviteins exhibited similar deltaG(degrees)(H(2)O) values of unfolding. Overall, it can be concluded that glycine caps are useful, as they reduce helical unravelling and enhance helicity, and thus, glycine will be included as a C-terminal residue in future de novo peptide sequences.


Assuntos
Éteres Cíclicos/química , Glicina/química , Peptídeos/química , Resorcinóis/química , Sequência de Aminoácidos , Dicroísmo Circular , Medição da Troca de Deutério , Dados de Sequência Molecular , Peptídeos/síntese química , Dobramento de Proteína , Estrutura Secundária de Proteína
4.
Biopolymers ; 90(1): 37-50, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-17994581

RESUMO

We investigated the hydrophobic packing of two previously designed caviteins, LG2 and LG3, which differ by one Gly in the linker regions between the peptide sequence and the cavitand scaffold. We sought to diminish the putative native-like properties of LG2 and LG3, and see if we could diagnose a change in the conformational specificity of the hydrophobic core. We replaced the leucine residues with norleucine residues at the hydrophobic positions in LG2 and LG3, to create NG2 and NG3, respectively. LG2 exhibited more dispersion, but less sharp signals than LG3 in the amide region of its (1)H NMR spectrum. NG3 and NG2 were found to be slightly less helical and significantly less stable toward guanidine hydrochloride compared with their reference caviteins. The (1)H NMR spectrum of NG2 was very similar to that of LG2, whereas there was a noticeable loss in the number and sharpness of the amide signals of NG3 compared with LG3. These data suggest that LG3 is very well packed; a loss in conformational specificity resulted from replacement of the leucine residues with norleucine residues. In contrast, the packing and dynamics of the hydrophobic core in LG2 were similar to those in NG2 (both more modest than LG3), as their (1)H NMR spectra were virtually indistinguishable. Overall, substitution of leucine by norleucine provided an efficient, convenient, and informative probe of the packing and dynamics of our caviteins' hydrophobic cores.


Assuntos
Éteres Cíclicos/química , Peptídeos/química , Resorcinóis/química , Dicroísmo Circular , Éteres Cíclicos/síntese química , Interações Hidrofóbicas e Hidrofílicas , Ressonância Magnética Nuclear Biomolecular , Peptídeos/síntese química , Proguanil/química , Desnaturação Proteica , Estrutura Secundária de Proteína , Espectrofotometria Ultravioleta
5.
Org Biomol Chem ; 5(22): 3637-50, 2007 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-17971993

RESUMO

The design, synthesis, and characterization of novel cavitand-based hetero-TASPs, TASPs having different peptide sequences within one bundle, are described. Three families of hetero-TASPs were designed: the LG3/LG2 family (different linker lengths), LG3/AG3 family (altering helix hydrophobicity), and the LG3/LG2C family (anti-parallel caviteins). These first generation hetero-TASPs were found to be alpha-helical, stable towards guanidine hydrochloride, and monomeric in solution. The LG3/LG2 caviteins exhibited primarily native-like properties. The remaining hetero-TASP families were found to exhibit less dispersion and broader signals in the amide regions of their (1)H NMR spectra than their respective reference caviteins. The success in the design of the LG3/LG2 hetero-TASPs suggests that subsequent hetero-TASPs may have potential to manifest superior native-like structure compared with homo-TASPs, and refinement of the linker and peptide sequences may accomplish this goal.


Assuntos
Éteres Cíclicos/química , Proteínas/química , Proteínas/síntese química , Resorcinóis/química , Amidas/química , Sequência de Aminoácidos , Dicroísmo Circular , Guanidina/farmacologia , Espectroscopia de Ressonância Magnética , Dados de Sequência Molecular , Peso Molecular , Naftalenossulfonatos/farmacologia , Peptídeos/química , Desnaturação Proteica/efeitos dos fármacos , Estrutura Secundária de Proteína , Prótons , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Ultracentrifugação
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