Probing the Membrane Targeting C1 Subdomains of PKC with Bivalent Ligands

Nowak, Ireneusz

doi:10.2174/156802607779941260

Cited by 4 publications

(3 citation statements)

References 40 publications

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“…First, what are the differential roles of the C1a and C1b domains of PKC␦ in ligand response? The existence of two C1 domains affords opportunities for complex modulation of PKC activity through bivalent ligands (35). However, although the performance of some of these ligands has shown improvement over that of monomers, it is inconsistent with expectations for cooperative binding to both C1 domains (21,35).…”

Section: Discussionmentioning

confidence: 99%

“…The existence of two C1 domains affords opportunities for complex modulation of PKC activity through bivalent ligands (35). However, although the performance of some of these ligands has shown improvement over that of monomers, it is inconsistent with expectations for cooperative binding to both C1 domains (21,35). Second, what are the relative contributions to ligand response of the C1 domain structure per se versus the position of the C1 domain within PKC␦?…”

Section: Discussionmentioning

confidence: 99%

“…Such studies provide clear examples that ligand-C1 domain interactions can be manipulated to yield novel patterns of recognition. Further selectivity might be gained with bivalent compounds, exploiting the spacing and individual characteristics of the C1a and C1b domains (35). A better understanding of the differential roles of the two C1 domains in PKC regulation is critical for the rational development of such compounds.…”

Section: H]pdbu Binding Membrane Translocation and Down-regulationmentioning

confidence: 99%

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Characterization of the Differential Roles of the Twin C1a and C1b Domains of Protein Kinase Cδ

Garfield

Kedei

et al. 2009

Journal of Biological Chemistry

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Classic and novel protein kinase C (PKC) isozymes contain two zinc finger motifs, designated "C1a" and "C1b" domains, which constitute the recognition modules for the second messenger diacylglycerol (DAG) or the phorbol esters. However, the individual contributions of these tandem C1 domains to PKC function and, reciprocally, the influence of protein context on their function remain uncertain. In the present study, we prepared PKC␦ constructs in which the individual C1a and C1b domains were deleted, swapped, or substituted for one another to explore these issues. As isolated fragments, both the ␦C1a and ␦C1b domains potently bound phorbol esters, but the binding of One of the essential steps for protein kinase C (PKC) 2 activation is its translocation from the cytosol to the membranes. For conventional (␣, ␤I, ␤II, and ␥) and novel (␦, ⑀, , and ) PKCs, this translocation is driven by interaction with the lipophilic second messenger sn-1,2-diacylglycerol (DAG), generated from phosphatidylinositol 4,5-bisphosphate upon the activation of receptor-coupled phospholipase C or indirectly from phosphatidylcholine via phospholipase D (1). A pair of zinc finger structures in the regulatory domain of the PKCs, the "C1" domains, are responsible for the recognition of the DAG signal. The DAG-C1 domain-membrane interaction is coupled to a conformational change in PKC, both causing the release of the pseudosubstrate domain from the catalytic site to activate the enzyme and triggering the translocation to the membrane (2). By regulating access to substrates, PKC translocation complements the intrinsic enzymatic specificity of PKC to determine its substrate profile.The C1 domain is a highly conserved cysteine-rich motif (ϳ50 amino acids), which was first identified in PKC as the interaction site for DAG or phorbol esters (3). It possesses a globular structure with a hydrophilic binding cleft at one end surrounded by hydrophobic residues. Binding of DAG or phorbol esters to the C1 domain caps the hydrophilic cleft and forms a continuous hydrophobic surface favoring the interaction or penetration of the C1 domain into the membrane (4). In addition to the novel and classic PKCs, six other families of proteins have also been identified, some of whose members possess DAG/phorbol ester-responsive C1 domains. These are the protein kinase D (5), the chimaerin (6), the munc-13 (7), the Ras-GRP (guanyl nucleotide exchange factors for Ras and Rap1) (8), the DAG kinase (9), and the recently characterized MRCK (myotonic dystrophy kinase-related Cdc42-binding kinase) families (10). Of these C1 domain-containing proteins, the PKCs have been studied most extensively and are important therapeutic targets (11). Among the drug candidates in clinical trials that target PKC, a number such as bryostatin 1 and PEP005 are directed at the C1 domains of PKC rather than at its catalytic site.Both the classic and novel PKCs contain in their N-terminal regulatory region tandem C1 domains, C1a and C1b, which bind DAG/phorbol ester (12). Multiple studies have...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: H]pdbu Binding Membrane Translocation and Down-regulationmentioning

confidence: 99%

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Characterization of the Differential Roles of the Twin C1a and C1b Domains of Protein Kinase Cδ

Garfield

Kedei

et al. 2009

Journal of Biological Chemistry

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Design and synthesis of a bivalent ligand to explore the putative heterodimerization of the mu opioid receptor and the chemokine receptor CCR5

et al. 2012

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The bivalent ligand approach has been utilized not only to study the underlying mechanism of G protein-coupled receptors dimerization and/or oligomerization, but also aimed to enhance ligand affinity and/or selectivity for potential treatment of a variety of diseases by targeting this process. Substance abuse and addiction have made both the prevention and the treatment of human immunodeficiency virus (HIV) infection more difficult to tackle. It has been extensively studied that morphine, a mu opioid receptor (MOR) agonist, can accelerate HIV infection through up-regulating the expression of the chemokine receptor CCR5, a well-known co-receptor for HIV invasion to the host cells. Meanwhile, two research groups have described the putative MOR/CCR5 heterodimers in their independent studies. The purpose of this paper is to report the design and synthesis of a bivalent ligand to explore the biological and pharmacological process of the putative MOR/CCR5 dimerization phenomenon. The developed bivalent ligand thus contains two distinct pharmacophores linked through a spacer; ideally one of which will interact with the MOR and the other with the CCR5. Naltrexone and Maraviroc were selected as the pharmacophores to generate such a bivalent probe. The overall reaction route to prepare this bivalent ligand was convergent and efficient, and involved sixteen steps with moderate to good yields. The preliminary biological characterization showed that the bivalent compound 1 retained the pharmacological characteristics of both pharmacophores towards the MOR and the CCR5 respectively with relatively lower binding affinity, which tentatively validated our original molecular design.

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A novel bivalent HIV-1 entry inhibitor reveals fundamental differences in CCR5-μ-opioid receptor interactions between human astroglia and microglia

El‐Hage¹,

Dever²,

Podhaizer³

et al. 2013

Aids

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Objective We explored whether the opiate, morphine, affects the actions of maraviroc, as well as a recently synthesized bivalent derivative of maraviroc linked to an opioid antagonist, naltrexone, on HIV-1 entry in primary human glia. Methods HIV-1 entry was monitored in glia transiently transfected with an LTR construct containing a luciferase reporter gene under control of a promoter for the HIV-1 transactivator protein Tat. The effect of maraviroc and the bivalent ligand ± morphine on CCR5 surface expression and cytokine release was also explored. Results Maraviroc inhibits HIV-1 entry into glial cells, while morphine negates the effects of maraviroc leading to a significant increase in viral entry. We also demonstrate that the maraviroc-containing bivalent ligand better inhibits R5-tropic viral entry in astrocytes than microglia compared to maraviroc when coadministered with morphine. Importantly, the inhibitory effects of the bivalent compound in astrocytes were not compromised by morphine. Exposure to maraviroc decreased the release of pro-inflammatory cytokines and restricted HIV-1-dependent increases in CCR5 expression in both astrocytes and microglia, while exposure to the bivalent had similar effect in astrocytes but not in microglia. CCR5-MOR stoichiometric ratio varied among the two cell types with CCR5 expressed at much higher levels than MOR in microglia, which could explain the effectiveness of the bivalent ligand in astrocytes compared to microglia. Conclusion A novel bivalent compound reveals fundamental differences in CCR5-MOR interactions and HIV-1 infectivity among glia, and has unique therapeutic potential in opiate abuse-HIV interactive comorbidity.

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Probing the Membrane Targeting C1 Subdomains of PKC with Bivalent Ligands

Cited by 4 publications

References 40 publications

Characterization of the Differential Roles of the Twin C1a and C1b Domains of Protein Kinase Cδ

Characterization of the Differential Roles of the Twin C1a and C1b Domains of Protein Kinase Cδ

Design and synthesis of a bivalent ligand to explore the putative heterodimerization of the mu opioid receptor and the chemokine receptor CCR5

A novel bivalent HIV-1 entry inhibitor reveals fundamental differences in CCR5-μ-opioid receptor interactions between human astroglia and microglia

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