Photoaffinity Labeling Analysis of the Interaction of Pituitary Adenylate‐Cyclase‐Activating Polypeptide (PACAP) with the PACAP Type I Receptor

Cao, Yong‐Jiang; Kojro, Elżbieta; Gimpl, Gerald; Jasionowski, Marek; Kasprzykowski, Franciszek; Łankiewicz, Leszek; Fahrenholz, Falk

doi:10.1111/j.1432-1033.1997.00400.x

Cited by 15 publications

(7 citation statements)

References 40 publications

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“…These results indicate that a charge reversing at cationic positions has a more deleterious effect on affinity than substituting these positions with Ala. Finally, incorporation of the photo-reactive amino acid benzoyl-phenylalanine at position 22 induces a 10-fold reduction of the affinity of PACAP27 [163]. Taken together, these results suggest that the helical segment of PACAP can tolerate different punctual modifications as long as they are not deleterious for the secondary structure of the peptide.…”

Section: Modifications Of the Central And C-terminal Segments Of Pacapmentioning

confidence: 75%

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Bourgault¹,

Vaudry²,

Dejda³

et al. 2009

CMC

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Pituitary adenylate cyclase-activating polypeptide (PACAP) is a 38-amino acid peptide that was initially isolated from hypothalamus extracts on the basis of its ability to stimulate the production of cAMP in cultured pituitary cells. Recent studies have shown that PACAP exerts potent neuroprotective effects not only in vitro but also in in vivo models of Parkinson's disease, Huntington's disease, traumatic brain injury and stroke. The protective effects of PACAP are based on its capacity to prevent neuronal apoptosis by acting directly on neurons or indirectly through the release of neuroprotective factors by astrocytes. These biological activities are mainly mediated through activation of the PAC1 receptor which is currently considered as a potential target for the treatment of neurodegenerative diseases. However, the use of native PACAP, the endogenous ligand of PAC1, as an efficient neuroprotective drug is actually limited by its rapid degradation. Moreover, injection of PACAP to human induces peripheral side effects which are mainly mediated through VPAC1 and VPAC2 receptors. Strategies to overcome these compromising conditions include the development of metabolically stable analogs of PACAP acting as selective agonists of the PAC1 receptor. This review presents an overview of the structure-activity relationships of PACAP and summarizes the molecular and conformational requirements for activation of PAC1 receptor. The applicability of PACAP analogs as therapeutic agents for treatment of neurodegenerative diseases is also discussed.

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Section: Modifications Of the Central And C-terminal Segments Of Pacapmentioning

confidence: 75%

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Bourgault¹,

Vaudry²,

Dejda³

et al. 2009

CMC

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“…This domain functions as the predominant pocket for binding the carboxyl-terminal regions of the natural ligands based on mutagenesis, ligand structure-activity, and photoaffinity labeling studies (3)(4)(5)(6)(7)(8)(9)(10)(11)(12). Recently, NMR and crystal structures have been described for segments of the isolated ligand-bound aminoterminal domain of the receptors for CRF 1 (13)(14)(15), gastric inhibitory polypeptide (16), GLP1 (17,18), pituitary adenylate cyclase activating peptide (19), PTH 1 (20), and calcitonin generelated peptide (21,22).…”

mentioning

confidence: 99%

Refinement of Glucagon-like Peptide 1 Docking to Its Intact Receptor Using Mid-region Photolabile Probes and Molecular Modeling

Miller

Chen

Lam

et al. 2011

Journal of Biological Chemistry

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The glucagon-like peptide 1 (GLP1) receptor is an important drug target within the B family of G protein-coupled receptors. Its natural agonist ligand, GLP1, has incretin-like actions and the receptor is a recognized target for management of type 2 diabetes mellitus. Despite recent solution of the structure of the amino terminus of the GLP1 receptor and several close family members, the molecular basis for GLP1 binding to and activation of the intact receptor remains unclear. We previously demonstrated molecular approximations between amino-and carboxyl-terminal residues of GLP1 and its receptor. In this work, we study spatial approximations with the mid-region of this peptide to gain insights into the orientation of the intact receptor and the ligand-receptor complex. We have prepared two new photolabile probes incorporating a p-benzoyl-L-phenylalanine into positions 16 and 20 of GLP1(7-36). Both probes bound to the GLP1 receptor specifically and with high affinity. These were each fully efficacious agonists, stimulating cAMP accumulation in receptor-bearing CHO cells in a concentration-dependent manner. Each probe specifically labeled a single receptor site. Protease cleavage and radiochemical sequencing identified receptor residue Leu 141 above transmembrane segment one as its site of labeling for the position 16 probe, whereas the position 20 probe labeled receptor residue Trp 297 within the second extracellular loop. Establishing ligand residue approximation with this loop region is unique among family members and may help to orient the receptor amino-terminal domain relative to its helical bundle region. G protein-coupled receptors (GPCRs)4 comprise a large family of plasma membrane receptors, binding molecules outside the cell that initiate intracellular signal transduction pathways and cellular responses. These molecules are targets for more than one-third of approved drugs. Family B GPCRs represent a relatively small group of receptors that nonetheless include important potential drug targets. Among these are receptors for secretin, vasoactive intestinal polypeptide, corticotropinreleasing factor (CRF), glucagon, glucagon-like peptide 1 (GLP1), calcitonin, and parathyroid hormone (PTH) (1).The GLP1 receptor is an important drug target within family B GPCRs. Its natural ligand, GLP1, has multiple antidiabetic actions, including promotion of insulin secretion, the preservation of ␤-cell mass, and the ability to lower body weight. A series of drugs have been developed and introduced that target this receptor for the treatment of type 2 diabetes mellitus (2). Like other natural ligands of this family, GLP1 is a moderately large and flexible peptide with a diffuse pharmacophore extending throughout its entire length, providing challenge to our understanding of its interaction with its receptor.Family B GPCRs all have a long and structurally characteristic extracellular amino-terminal domain stabilized by three pairs of disulfide bonds linking six conserved cysteine residues. This domain functions as the pred...

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“…Photoaffinity labeling of the PACAP receptor was performed as described previously. 29 Membranes (100 μg) were incubated with 0.5 nM each of the iodinated photoreactive analogues ( 125 I‐[Phe 6 (pBz)]PACAP(1‐27), 125 I‐[Phe 22 (pBz)]PACAP(1‐27), 125 I‐[Lys 15 (ɛ‐pBz 2 )]PACAP(1‐27), 125 I‐[Lys 20 (ɛ‐pBz 2 )]PACAP(1‐27), and 125 I‐[Lys 21 (ɛ‐pBz 2 )]PACAP(1‐27)) in a total volume of 200 μl binding buffer B (buffer A without BSA) for 60 min at 25°C. The reaction was then quenched by either diluting the mixture in 2 ml binding buffer B or by pelleting the membrane and resuspending it in 1 ml of the same buffer.…”

Section: Methodsmentioning

confidence: 99%

“…Photoaffinity labeling of the PACAP receptor was performed as described previously. 29 Membranes (100 µg) were incubated with 0. in a total volume of 200 µl binding buffer B (buffer A without BSA) for 60 min at 25°C. The reaction was then quenched by either diluting the mixture in 2 ml binding buffer B or by pelleting the membrane and resuspending it in 1 ml of the same buffer.…”

Section: Photoaffinity Labeling Of Pacap Receptorsmentioning

confidence: 99%

Identification of Binding Domains of Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) for its Type 1 Receptor by Photoaffinity Labeling^a

Cao

Kojro

Jasionowski

et al. 1998

Annals of the New York Academy of Sciences

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Structure-function studies and photoaffinity labeling experiments were performed to identify residues and domains of PACAP involved in the interaction with PACAP receptors. For this purpose, a series of photoreactive analogues of PACAP(1-27) containing a photoreactive benzophenone (BP) residue in different peptide structural domains were utilized to analyze the interaction of PACAP(1-27) with pig PACAP type 1 receptors. Five PACAP derivatives were created with a photoreactive amino acid in the following peptide domains: either the disordered N-terminal or the helical C-terminal domain or a short loop region within the C-terminal helical domain of the peptide. Their receptor binding properties and efficiencies were tested on pig brain PACAP receptors. The results indicate the importance of the helical C-terminal domain of PACAP(1-27) for receptor binding affinity. Monoiodination of the photoreactive analogues did not change their binding affinities. Experiments with pig brain membranes demonstrated that the 125I-labeled photoreactive analogues specifically label a protein band of M(r) 66,000. The efficiency of photoreactive labeling differed for the various analogues. These findings suggest that Tyr22 and Lys15 in PACAP (1-27) are located in or close to the hormone binding site of the PACAP type 1 receptor. The results provide evidence that the alpha-helical C-terminal region of PACAP is directly involved in receptor binding.

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Photoaffinity Labeling Analysis of the Interaction of Pituitary Adenylate‐Cyclase‐Activating Polypeptide (PACAP) with the PACAP Type I Receptor

Cited by 15 publications

References 40 publications

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Refinement of Glucagon-like Peptide 1 Docking to Its Intact Receptor Using Mid-region Photolabile Probes and Molecular Modeling

Identification of Binding Domains of Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) for its Type 1 Receptor by Photoaffinity Labeling^a

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Photoaffinity Labeling Analysis of the Interaction of Pituitary Adenylate‐Cyclase‐Activating Polypeptide (PACAP) with the PACAP Type I Receptor

Cited by 15 publications

References 40 publications

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Pituitary Adenylate Cyclase-Activating Polypeptide: Focus on Structure- Activity Relationships of a Neuroprotective Peptide

Refinement of Glucagon-like Peptide 1 Docking to Its Intact Receptor Using Mid-region Photolabile Probes and Molecular Modeling

Identification of Binding Domains of Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) for its Type 1 Receptor by Photoaffinity Labelinga

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Identification of Binding Domains of Pituitary Adenylate Cyclase Activating Polypeptide (PACAP) for its Type 1 Receptor by Photoaffinity Labeling^a