Mimotopes: realization of an unlikely concept

Meloen, Rob H.; Puijk, Wouter C.; Slootstra, Jerry W.

doi:10.1002/1099-1352(200011/12)13:6<352::aid-jmr509>3.0.co;2-c

Cited by 88 publications

(43 citation statements)

References 71 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Several other tools and methods have been developed to study protein-protein interactions, such as multi-use peptide libraries (8) and spot synthesis (9, 10), alanine (11,12) or cysteine (38) scanning, and two-hybrid analysis (39,40). Our technique has important advantages over these methods.…”

Section: Discussionmentioning

confidence: 99%

“…Probing the accessible surface in protein complexes by deuterium exchange has also been employed (7). But by far the two most popular methods are the use of peptide mimetics either in solution (8) or on membranes (9,10) and especially site-directed mutagenesis (11,12). However, in the latter powerful technique to study functions of proteins or interactions between proteins, one often faces the difficulty of choosing the position and type of amino acid exchange to be introduced.…”

mentioning

confidence: 99%

See 1 more Smart Citation

A Phage Display-based Method for Determination of Relative Affinities of Mutants

Rossenu

Leyman²,

Dewitte

et al. 2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

We propose phage display combined with enzymelinked immunosorbent assay as a tool for the systematic analysis of protein-protein interactions by investigating the binding behavior of variants to a partner protein. Via enzyme-linked immunosorbent assay we determine both the amount of fusion protein presented at the phage surface and the amount of complex formed, the ratio of which is proportional to the affinity. Hence this method enables us to calculate the relative affinities of a large number of mutants. As model systems, we investigated actin-binding motifs conserved in a number of proteins binding monomeric or filamentous actin. The hexapeptide motifs LKKTET, present in thymosin ␤4, and LKKEKG, present in the villin headpiece, were mutated, and the variants were analyzed. Study of the positional tolerance allows postulating that the motifs, although similar in primary structures adopt different conformations when bound to actin. In addition, our data show that the second and the fourth amino acid of the thymosin ␤4 motif and the first three residues of the villin headpiece motif are most important for actin binding. The latter result challenges the charged crown hypothesis for the villin headpiece filamentous actin interaction.As a consequence of various genome-sequencing projects, novel proteins are being identified, many of which may take part in new interactions. Efforts are going on to tackle the discovery of protein-protein interactions globally (1), but there is also need for novel methods for the analysis of these interactions, preferably easy, reliable, and high through-put techniques. Several means to probe amino acids that contribute to the binding of two proteins have been developed. X-ray crystallography and NMR may yield superior information about the interface of proteins at atomic resolution (2, 3). However, both methods are intrinsically difficult (even for noncomplexed proteins) and time-consuming and require expensive and sophisticated equipment, as well as large amounts of biological material. Cross-linking of interacting proteins followed by mass spectroscopy or conventional sequence determination of covalently coupled peptide fragments has limited application (4 -6). Probing the accessible surface in protein complexes by deuterium exchange has also been employed (7). But by far the two most popular methods are the use of peptide mimetics either in solution (8) or on membranes (9, 10) and especially site-directed mutagenesis (11, 12). However, in the latter powerful technique to study functions of proteins or interactions between proteins, one often faces the difficulty of choosing the position and type of amino acid exchange to be introduced. To circumvent this, one may perform a saturation mutagenesis at several positions thought to be important for the interaction. Obviously this creates a new problem, i.e. screening a large number of mutants in a systematic way.We propose to combine saturation mutagenesis, phage display and ELISA 1 for the systematic screening and quantification of pr...

show abstract

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

A Phage Display-based Method for Determination of Relative Affinities of Mutants

Rossenu

Leyman²,

Dewitte

et al. 2003

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Selective agonists or antagonists of these natural peptides are extremely useful for the investigation of peptidergic systems and are also potential therapeutic agents (1). Moreover, several peptide fragments or mimotopes derived from potential therapeutic proteins show promising biological activity (2).…”

mentioning

confidence: 99%

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

Bracci

Falciani

Lelli

et al. 2003

Journal of Biological Chemistry

156

162

View full text Add to dashboard Cite

In previous papers, we observed that dendrimers of peptide mimotopes of the nicotinic receptor ligand site are strong antidotes against the lethality of the nicotinic receptor ligand ␣-bungarotoxin. Although their in vitro activity is identical to that of dendrimers, the corresponding monomeric peptide mimotopes are not effective in vivo. Because the higher in vivo efficiency of dendrimers could not in this case be related to polyvalent interaction, the stability to blood protease activity of monomeric versus tetrabranched dendrimeric mimotope peptides was compared here by incubating three different mimotopes with human plasma and serum. Unmodified peptides and cleaved sequences were followed by high pressure liquid chromatography and mass spectrometry. Tetrabranched peptides were shown to be much more stable in plasma and also in serum. To assess the notable stability of multimeric peptides, different bioactive neuropeptides, including enkephalins, neurotensin and nociceptin, were synthesized in monomeric and tetrabranched forms and incubated with human plasma and serum and with rat brain membrane extracts. All the tetrabranched neuropeptides fully retained biological activity and generally showed much greater stability to blood and brain protease activity. Some tetrabranched peptides were also resistant to trypsin and chymotrypsin. Our findings provide new insights into the possible therapeutic use of bioactive peptides.Hundreds of peptides with potential therapeutic activities have been identified. These include naturally occurring peptide hormones and neurotransmitters, which influence and control series of vital functions, such as cell proliferation, tissue development, metabolism, immune defense, perception of pain, reproduction, behavior, and blood pressure. Selective agonists or antagonists of these natural peptides are extremely useful for the investigation of peptidergic systems and are also potential therapeutic agents (1). Moreover, several peptide fragments or mimotopes derived from potential therapeutic proteins show promising biological activity (2).However, the use of peptides as therapeutic drugs has largely been limited by their short half-life in vivo. Because peptides are mainly broken down by proteases and peptidases, peptide delivery is the bottleneck in the development of new peptide drugs. To increase peptide half-life, many strategies involving different levels of chemical modification are possible (3, 4). The introduction of D-amino acids, or pseudo amino acids, and peptide cyclization are the most common strategies to increase peptide stability. However, these modifications may profoundly alter peptide activity. Alternatively, peptidomimetic molecules can be developed by the synthesis of conformationally restricted compounds, in which the peptide is locally or globally constrained in order to reproduce the active conformation. The resulting structures are mostly non-peptide molecules, more resistant to degrading enzymes.In general, peptide molecules have the advantage of good specifici...

show abstract

“…Molecular mimicry to the cognate antigen seems to be required for inducing a functional, cross-reactive immune response. 1 In several studies this functional molecular mimicry has been achieved by a trial-and-error approach, 2 -7 by combinatorial chemistry display techniques 8,9 or by structure-based design. 10 -12 Similar to approaches in structure-based drug design, crystal or solution structures of peptide-antibody complexes are often used to optimize the complementarity between a peptide and a functional antibody, which should thus lead to higher binding affinity.…”

mentioning

confidence: 99%

Immunogenicity of Peptide-vaccine Candidates Predicted by Molecular Dynamics Simulations

Oomen

Hoogerhout

Bonvin

et al. 2003

Journal of Molecular Biology

View full text Add to dashboard Cite

We present an in silico, structure-based approach for design and evaluation of conformationally restricted peptide-vaccines. In particular, we designed four cyclic peptides of ten or 11 residues mimicking the crystallographically observed b-turn conformation of a predicted immunodominant loop of PorA from Neisseria meningitidis. Conformational correctness and stability of the peptide designs, as evaluated by molecular dynamics simulations, correctly predicted the immunogenicity of the peptides. We observed a peptide-induced functional antibody response that, remarkably, exceeded the response induced by the native protein in outer membrane vesicles, without losing specificity for related strains. The presented approach offers tools for a priori design and selection of peptide-vaccine candidates with full biological activity. This approach could be widely applicable: to outer membrane proteins of Gram-negative bacteria, and to other epitopes in a large range of pathogens.

show abstract

Mimotopes: realization of an unlikely concept

Cited by 88 publications

References 71 publications

A Phage Display-based Method for Determination of Relative Affinities of Mutants

A Phage Display-based Method for Determination of Relative Affinities of Mutants

Synthetic Peptides in the Form of Dendrimers Become Resistant to Protease Activity

Immunogenicity of Peptide-vaccine Candidates Predicted by Molecular Dynamics Simulations

Contact Info

Product

Resources

About