ABSTRACT
The ligands of certain G-protein-coupled receptors (GPCRs) have been identified as endogenous lipids, such as lysophosphatidylserine (LysoPS). Here, we analyzed the molecular basis of the structure-activity relationship of ligands of GPR34, one of the LysoPS receptor subtypes, focusing on recognition of the long-chain fatty acid moiety by the hydrophobic pocket. By introducing benzene ring(s) into the fatty acid moiety of 2-deoxy-LysoPS, we explored the binding site's preference for the hydrophobic shape. A tribenzene-containing fatty acid surrogate with modifications of the terminal aromatic moiety showed potent agonistic activity toward GPR34. Computational docking of these derivatives with a homology modeling/molecular dynamics-based virtual binding site of GPR34 indicated that a kink in the benzene-based lipid surrogates matches the L-shaped hydrophobic pocket of GPR34. A tetrabenzene-based lipid analogue bearing a bulky tert-butyl group at the 4-position of the terminal benzene ring exhibited potent GPR34 agonistic activity, validating the present hydrophobic binding pocket model.
Subject(s)
Benzene Derivatives/chemistry , Fatty Acids/chemistry , Phosphoserine/analogs & derivatives , Receptors, Lysophospholipid/chemistry , Animals , Benzene Derivatives/chemical synthesis , Benzene Derivatives/pharmacology , Binding Sites , Fatty Acids/chemical synthesis , Fatty Acids/pharmacology , HEK293 Cells , Humans , Hydrophobic and Hydrophilic Interactions , Mice , Molecular Docking Simulation , Molecular Dynamics Simulation , Phosphoserine/chemical synthesis , Phosphoserine/chemistry , Phosphoserine/pharmacology , Receptors, Lysophospholipid/agonists , Structure-Activity RelationshipABSTRACT
This overview focuses on the (alpha,alpha-difluoromethylene)phosphonate mimic of phosphoserine (pCF(2)Ser) and its application to the study of kinase-mediated signal transduction-pathways of great interest to drug development. The most versatile modes of access to these chemical biological tools are discussed, organized by method of PCF(2)-C bond formation. The pCF(2)-Ser mimic may be site-specifically incorporated into peptides (SPPS) and proteins (expressed protein ligation). This isopolar, dianionic pSer mimic results in a "constitutive phosphorylation" phenotype and is seen to support native protein-protein interactions that depend on serine phosphorylation. Signal transduction pathways studied with this chemical biological approach include the regulation of p53 tumor suppressor protein activity and of melatonin production. Given these successes, the future is bright for the use of such "teflon phospho-amino acid mimics" to map kinase-based signaling pathways.
Subject(s)
Clodronic Acid/analogs & derivatives , Phosphoserine/chemistry , Signal Transduction , Clodronic Acid/chemical synthesis , Clodronic Acid/chemistry , Clodronic Acid/pharmacology , MAP Kinase Signaling System , Melatonin/metabolism , Phosphorylation , Phosphoserine/chemical synthesis , Phosphoserine/pharmacology , Protein Interaction Domains and Motifs , Tumor Suppressor Protein p53/metabolismABSTRACT
[reaction: see text] An interassembly approach for the synthesis of peptides containing 1-(2-nitrophenyl)ethyl-caged phosphoserine, -threonine, and -tyrosine has been developed. Photochemical uncaging of these peptides releases the 2-nitrophenylethyl protecting group to afford the corresponding phosphopeptide. The peptides described herein are based on phosphorylation sites of kinases involved in cell movement or cell cycle regulation and demonstrate the versatility of the method and compatibility with the synthesis of polypeptides, including a variety of encoded amino acids.
Subject(s)
Peptides/chemical synthesis , Phosphorylation , Phosphoserine/chemical synthesis , Phosphothreonine/chemical synthesis , Phosphotyrosine/chemical synthesis , Photolysis , Signal TransductionABSTRACT
An improved synthesis of two lipid phosphoric acids, N-palmitoyl-L-serine phosphoric acid (NP-Ser-PA) and N-palmitoyl-L-tyrosine phosphoric acid (NP-Tyr-PA), from the benzyl esters of L-serine and L-tyrosine is described. The sequence of N-acylation, followed by phosphitylation with N, N-diisopropyl dibenzyl phosphoramidite, oxidation to the corresponding phosphate triesters, and simultaneous debenzylation of the dibenzyl phosphate and benzyl carboxylic esters gave NP-Ser-PA and NP-Tyr-PA in high overall yields. NP-Ser-PA and NP-Tyr-PA and their D stereoisomers were potent reversible inhibitors of the lysophosphatidic acid receptors expressed in Xenopus oocytes, thus providing prototypic structures for the development of inhibitors of the lysophosphatidate family of phospholipid growth factors.
Subject(s)
Palmitates/chemical synthesis , Phosphoserine/analogs & derivatives , Phosphotyrosine/analogs & derivatives , Receptors, Cell Surface/antagonists & inhibitors , Receptors, G-Protein-Coupled , Animals , Cell Membrane/physiology , Chloride Channels/drug effects , Chloride Channels/physiology , Electric Conductivity , Female , Oocytes/metabolism , Oocytes/ultrastructure , Palmitates/pharmacology , Phosphoserine/chemical synthesis , Phosphoserine/pharmacology , Phosphotyrosine/chemical synthesis , Phosphotyrosine/pharmacology , Receptors, Lysophosphatidic Acid , Recombinant Proteins/antagonists & inhibitors , Stereoisomerism , XenopusSubject(s)
Molecular Biology/methods , Phosphopeptides/chemical synthesis , Amino Acid Sequence , Indicators and Reagents , Molecular Sequence Data , Phosphopeptides/chemistry , Phosphopeptides/isolation & purification , Phosphorylation , Phosphoserine/chemical synthesis , Phosphoserine/chemistry , Phosphothreonine/chemical synthesis , Phosphothreonine/chemistryABSTRACT
The multiple Ser(P)-containing peptides, H-Ser(P)-Ser(P)-Ser(P)-Glu-Glu-NHMe.TFA, H-Asp-Ser(P)-Ser(P)-Glu-Glu-NHMe.TFA and H-Glu-Ser(P)-Ser(P)-Glu-Glu-NHMe.TFA were prepared by the use of Boc-Ser(PO3Ph2)-OH in the Boc mode of solution phase peptide synthesis followed by platinum-mediated hydrogenolytic de-protection of the Ser(PO3Ph2)-containing peptides. The protected peptides were assembled using the mixed anhydride coupling methods with 40% TFA/CH2Cl2 used for removal of the Boc group from intermediate Boc-protected peptides.
