Biologically displayed random peptides as reagents in mapping protein-protein interactions

Kay, Brian K.

doi:10.1007/bf02172066

Cited by 23 publications

(8 citation statements)

References 120 publications

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“…This most likely explains why a small peptide, like pT5, requires a high concentration to impair the troponin activation in situ. Taken together, these results demonstrate the power of using phage-displayed combinatorial libraries to map native protein-protein interactions (3,7). The results also implicate that this short N-terminal sequence within TnI plays an important role in the interaction of TnC and TnI in vivo, which is supported by the results of previous studies on the interaction of TnC and TnI.…”

Section: Discussionsupporting

confidence: 81%

Identification of Troponin C Antagonists from a Phage-displayed Random Peptide Library

Pierce¹,

Schachat²,

Brandt³

et al. 1998

Journal of Biological Chemistry

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Affinity purification of a phage-displayed library, expressing random peptide 12-mers at the N terminus of protein III, has identified 10 distinct novel sequences which bind troponin C specifically. The troponin C-selected peptides yield a consensus binding sequence of (V/L)(D/E)XLKXXLXXLA. Sequence comparison revealed as much as a 62.5% similarity between T5, the peptide sequence of the phage clone with the highest level of binding to troponin C, and the N-terminal region of troponin I isoforms. Biotinylated peptides corresponding to library-derived sequences and similar sequences from various isoforms of troponin I were synthesized shown to bind troponin C specifically. Alkaline phosphatase fusion proteins of two of the phage clone sequences bound troponin C specifically, and were specifically competed by both library-derived and native troponin I peptides. Measurement of equilibrium dissociation constants of the peptides by surface plasmon resonance yielded dissociation constants for troponin C as low as 0.43 M for pT5; in contrast, dissociation constants for calmodulin were greater than 6 M for all peptides studied. Nondenaturing polyacrylamide gel electrophoresis demonstrated that pT5 formed a stable complex with troponin C in the presence of calcium. We also found that the pT5 peptide inhibited the maximal calcium-activated tension of rabbit psoas muscle fibers.

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Section: Discussionsupporting

confidence: 81%

Identification of Troponin C Antagonists from a Phage-displayed Random Peptide Library

Pierce¹,

Schachat²,

Brandt³

et al. 1998

Journal of Biological Chemistry

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“…Because peptide displayed by phage libraries has been widely used to infer protein ligands of target molecules,22, 23 we searched the databases for similarity with the high‐affinity peptide sequences 31. Conventional BLASTP search on E. coli database did not retrieve either known interacting partners of FtsA, nor other proteins involved in cell division.…”

Section: Resultsmentioning

confidence: 99%

“…We chose to investigate the binding features of FtsA using affinity screening of a peptide library displayed on the surface of M13 bacteriophages. This powerful technique has been widely applied to develop receptor‐specific ligands, drug‐delivery systems, and enzyme inhibitors and substrates 22, 23. Concerning FtsA, the identification of specific peptide ligands appeared attractive both to shed light on its poorly defined binding specificity and to explore their use as templates to discover novel antimicrobial lead compounds with broad spectrum activity.…”

Section: Introductionmentioning

confidence: 99%

Phage‐display and correlated mutations identify an essential region of subdomain 1C involved in homodimerization of Escherichia coli FtsA

et al. 2002

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FtsA plays an essential role in Escherichia coli cell division and is nearly ubiquitous in eubacteria. Several evidences postulated the ability of FtsA to interact with other septation proteins and with itself. To investigate these binding properties, we screened a phage-display library with FtsA. The isolated peptides defined a degenerate consensus sequence, which in turn displayed a striking similarity with residues 126-133 of FtsA itself. This result suggested that residues 126-133 were involved in homodimerization of FtsA. The hypothesis was supported by the analysis of correlated mutations, which identified a mutual relationship between a group of amino acids encompassing the ATP-binding site and a set of residues immediately downstream to amino acids 126-133. This information was used to assemble a model of a FtsA homodimer, whose accuracy was confirmed by probing multiple alternative docking solutions. Moreover, a prediction of residues responsible for protein-protein interaction validated the proposed model and confirmed once more the importance of residues 126-133 for homodimerization. To functionally characterize this region, we introduced a deletion in ftsA, where residues 126-133 were skipped. This mutant failed to complement conditional lethal alleles of ftsA, demonstrating that amino acids 126-133 play an essential role in E. coli.

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“…The terms in Eqns. (13) and (15) and the rst four terms in Eqns. (12) and (14) are the standard ows to and from free, singly-bound and doubly-bound states.…”

Section: Modelmentioning

confidence: 99%

Stochastic modeling and optimization of phage display 1 1Edited by J. A. Wells

Levitan

1998

Journal of Molecular Biology

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Biologically displayed random peptides as reagents in mapping protein-protein interactions

Cited by 23 publications

References 120 publications

Identification of Troponin C Antagonists from a Phage-displayed Random Peptide Library

Identification of Troponin C Antagonists from a Phage-displayed Random Peptide Library

Phage‐display and correlated mutations identify an essential region of subdomain 1C involved in homodimerization of Escherichia coli FtsA

Stochastic modeling and optimization of phage display 1 1Edited by J. A. Wells

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