Rapid Affinity‐Based Fingerprinting of 14‐3‐3 Isoforms Using a Combinatorial Peptide Microarray

Lu, Candy H. S.; Sun, Hongyan; Bakar, Farhana B. Abu; Uttamchandani, Mahesh; Zhou, Wei; Liou, Yih‐Cherng; Yao, Shao Q.

doi:10.1002/anie.200801395

Cited by 35 publications

(19 citation statements)

References 23 publications

(28 reference statements)

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“…[7] We therefore first determined the most preferred pSer-containing heptapeptide that binds to PARP1 BRCT by screening the fluorescently labeled recombinant protein with a previously reported, 1000-member universal phosphopeptide microarray (Step I). [16] We determined the most preferred PARP1 BRCT domain binding peptide as LRFpSVFF (Figure 2 b), which, aside from the pSer residue, contains a P +3 Phe residue as the other major binding determinant. Next, we further optimized this sequence by constructing a 115-member positional scanning (PS) heptapeptide microarray (Step II); all six residues in LRFpSVFF (except pS) were individually substituted with each of the 20 naturally occurring amino acids.…”

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confidence: 99%

A Small‐Molecule Protein–Protein Interaction Inhibitor of PARP1 That Targets Its BRCT Domain

Peng

et al. 2015

Angew Chem Int Ed

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Poly(ADP-ribose)polymerase-1 (PARP1) is a BRCT-containing enzyme (BRCT = BRCA1 C-terminus) mainly involved in DNA repair and damage response and a validated target for cancer treatment. Small-molecule inhibitors that target the PARP1 catalytic domain have been actively pursued as anticancer drugs, but are potentially problematic owing to a lack of selectivity. Compounds that are capable of disrupting protein-protein interactions of PARP1 provide an alternative by inhibiting its activities with improved selectivity profiles. Herein, by establishing a high-throughput microplate-based assay suitable for screening potential PPI inhibitors of the PARP1 BRCT domain, we have discovered that (±)-gossypol, a natural product with a number of known biological activities, possesses novel PARP1 inhibitory activity both in vitro and in cancer cells and presumably acts through disruption of protein-protein interactions. As the first known cell-permeable small-molecule PPI inhibitor of PAPR1, we further established that (-)-gossypol was likely the causative agent of PARP1 inhibition by promoting the formation of a 1:2 compound/PARP1 complex by reversible formation of a covalent imine linkage.

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confidence: 99%

A Small‐Molecule Protein–Protein Interaction Inhibitor of PARP1 That Targets Its BRCT Domain

Peng

et al. 2015

Angew Chem Int Ed

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“…[69] We recently extended this strategy by reporting what we called a fragment-based combinatorial peptide library consisting of 1000 different spotting features on a single glass slide (Figure 6; workflow II). [70] With this new platform, a much larger peptide sequence space could be covered with a manageable number of spots/peptide libraries, without the loss of detailed and subtle information on protein-peptide interactions. Subsequent high-throughput screening was carried out to determine the substrate binding specificity of the highly homolowww.chemeurj.org gous 14-3-3 proteins (a class of PBDs).…”

Section: Biosensors That Report Phosphorylation Eventsmentioning

confidence: 99%

Current Chemical Biology Tools for Studying Protein Phosphorylation and Dephosphorylation

Tan

et al. 2011

Chemistry A European J

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Amongst different posttranslational events involved in cellular-signaling pathways, phosphorylation and dephosphorylation of proteins are the most prevalent. Aberrant regulations in the cellular phosphoproteome network are implicated in most major human diseases. Consequently, kinases and phosphatases are two of the most important groups of drug targets in medicinal research today. A major challenge in the understanding of protein phosphorylation and dephosphorylation is the sheer complexity of the phosphoproteome network and the lack of tools capable of studying protein phosphorylation and dephosphorylation as they occur in cells. We highlight herein various chemical biology tools that have emerged in the last decade for such studies. First, we discuss the use of small-molecule mimics of phosphoamino acids and their use in elucidating the function of protein phosphorylation and dephosphorylation. We also introduce recent advances in the field of activity-based protein profiling (ABPP) for proteome-wide detection of protein phosphorylation and dephosphorylation. We next discuss the key concepts in the design of peptide- and protein-based biosensors capable of real-time reporting of phosphorylation/dephosphorylation events. Finally, we highlight the application of peptide and small-molecule microarrays (SMMs), and their applications in high-throughput screening and discovery of new compounds related to phosphorylation/dephosphorylation.

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“…A 1000-member fragment-based phosphoserine/threonine heptapeptide library was generated to screen the activity of the family of seven human 14-3-3 proteins on microarrays, which are important cellular regulators that bind to phosphoserine-containing proteins. 118 14-3-3 proteins are very closely related, and it has been a challenge to identify their unique target specificities. The microarray-based two-color screening method revealed a novel target sequence that bound 14-3-3 Sigma, which is a protein implicated in tumorigenesis.…”

Section: B Inhibitor Profiling and Protein Fingerprintingmentioning

confidence: 99%

“…The microarray-based two-color screening method revealed a novel target sequence that bound 14-3-3 Sigma, which is a protein implicated in tumorigenesis. 118 In another interesting development, Dordick et al developed cytochrome P450 ͑CYP͒ arrays that are able to screen for enzyme activity and inhibition. In one demonstration, sol-gel encapsulated P450 enzymes on microarrays were used to activate a prodrug, cyclophosphamide.…”

Section: B Inhibitor Profiling and Protein Fingerprintingmentioning

confidence: 99%

Microarray-based enzyme profiling: Recent advances and applications (Review)

Uttamchandani

Moochhala

2010

Biointerphases

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Enzymes are an integral part of biological systems. They constitute a significant majority of all proteins expressed (an estimated 18%–29%) within eukaryotic genomes. It thus comes as no major surprise that enzymes have been implicated in many diseases and form the second largest group of drug targets, after receptors. Despite their involvement in a multitude of physiological processes, only a limited number of enzymes have thus far been well-characterized. Consequently, little is understood about the physiological roles, substrate specificity, and downstream targets of the vast majority of these important proteins. In order to facilitate the biological characterization of enzymes, as well as their adoption as drug targets, there is a need for global “-omics” solutions that bridge the gap in understanding these proteins and their interactions. Herein the authors showcase how microarray methods can be adopted to facilitate investigations into enzymes and their properties, in a high-throughput manner. They will focus on several major classes of enzymes, including kinases, phosphatases, and proteases. As a result of research efforts over the last decade, these groups of enzymes have become readily amenable to microarray-based profiling methods. The authors will also describe the specific design considerations that are required to develop the appropriate chemical tools and libraries to characterize each enzyme class. These include peptide substrates, activity-based probes, and chemical compound libraries, which may be rapidly assembled using efficient combinatorial synthesis or “click chemistry” strategies. Taken together, microarrays offer a powerful means to study, profile, and also discover potent small molecules with which to modulate enzyme activity.

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Rapid Affinity‐Based Fingerprinting of 14‐3‐3 Isoforms Using a Combinatorial Peptide Microarray

Cited by 35 publications

References 23 publications

A Small‐Molecule Protein–Protein Interaction Inhibitor of PARP1 That Targets Its BRCT Domain

A Small‐Molecule Protein–Protein Interaction Inhibitor of PARP1 That Targets Its BRCT Domain

Current Chemical Biology Tools for Studying Protein Phosphorylation and Dephosphorylation

Microarray-based enzyme profiling: Recent advances and applications (Review)

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