Glucagon-like peptide-1 synthetic analogs: new therapeutic agents for use in the treatment of diabetes mellitus.

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Glucagon-like peptide-1 synthetic analogs: new therapeutic agents for use in the treatment of diabetes mellitus.

Curr Med Chem. 2003 Nov;10(22):2471-83

Authors: Holz GG, Chepurny OG

Abstract
Glucagon-like peptide-1-(7-36)-amide (GLP-1) is a potent blood glucose-lowering hormone now under investigation for use as a therapeutic agent in the treatment of type 2 (adult onset) diabetes mellitus. GLP-1 binds with high affinity to G protein-coupled receptors (GPCRs) located on pancreatic beta-cells, and it exerts insulinotropic actions that include the stimulation of insulin gene transcription, insulin biosynthesis, and insulin secretion. The beneficial therapeutic action of GLP-1 also includes its ability to act as a growth factor, stimulating formation of new pancreatic islets (neogenesis) while slowing beta-cell death (apoptosis). GLP-1 belongs to a large family of structurally-related hormones and neuropeptides that include glucagon, secretin, GIP, PACAP, and VIP. Biosynthesis of GLP-1 occurs in the enteroendocrine L-cells of the distal intestine, and the release of GLP-1 into the systemic circulation accompanies ingestion of a meal. Although GLP-1 is inactivated rapidly by dipeptidyl peptidase IV (DDP-IV), synthetic analogs of GLP-1 exist, and efforts have been directed at engineering these peptides so that they are resistant to enzymatic hydrolysis. Additional modifications of GLP-1 incorporate fatty acylation and drug affinity complex (DAC) technology to improve serum albumin binding, thereby slowing renal clearance of the peptides. NN2211, LY315902, LY307161, and CJC-1131 are GLP-1 synthetic analogs that reproduce many of the biological actions of GLP-1, but with a prolonged duration of action. AC2993 (Exendin-4) is a naturally occurring peptide isolated from the lizard Heloderma, and it acts as a high affinity agonist at the GLP-1 receptor. This review summarizes structural features and signal transduction properties of GLP-1 and its cognate beta-cell GPCR. The usefulness of synthetic GLP-1 analogs as blood glucose-lowering agents is discussed, and the applicability of GLP-1 as a therapeutic agent for treatment of type 2 diabetes is highlighted.

PMID: 14529486 [PubMed - indexed for MEDLINE]

NF-kB pathway NF-kB signaling NF-kappaB signaling pathway

Chimeric yeast G-protein ? subunit harboring a 37-residue C-terminal gustducin-specific sequence is functional in Saccharomyces cerevisiae.

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Chimeric yeast G-protein ? subunit harboring a 37-residue C-terminal gustducin-specific sequence is functional in Saccharomyces cerevisiae.

Biosci Biotechnol Biochem. 2012;76(3):512-6

Authors: Hara K, Inada Y, Ono T, Kuroda K, Yasuda-Kamatani Y, Ishiguro M, Tanaka T, Misaka T, Abe K, Ueda M

Abstract
Despite many recent studies of G-protein-coupled receptor (GPCR) structures, it is not yet well understood how these receptors activate G proteins. The GPCR assay using baker's yeast, Saccharomyces cerevisiae, is an effective experimental model for the characterization of GPCR-G? interactions. Here, using the yeast endogenous G? protein (Gpa1p) as template, we constructed various chimeric G? proteins with a region that is considered to be necessary for interaction with mammalian receptors. The signaling assay using the yeast pheromone receptor revealed that the chimeric G? protein harboring 37 gustducin-specific amino acid residues at its C-terminus (GPA1/gust37) maintained functionality in yeast. In contrast, GPA1/gust44, a variant routinely used in mammalian experimental systems, was not functional.

PMID: 22451393 [PubMed - indexed for MEDLINE]

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Modifying ligand-induced and constitutive signaling of the human 5-HT4 receptor.

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Modifying ligand-induced and constitutive signaling of the human 5-HT4 receptor.

PLoS One. 2007;2(12):e1317

Authors: Chang WC, Ng JK, Nguyen T, Pellissier L, Claeysen S, Hsiao EC, Conklin BR

Abstract
G protein-coupled receptors (GPCRs) signal through a limited number of G-protein pathways and play crucial roles in many biological processes. Studies of their in vivo functions have been hampered by the molecular and functional diversity of GPCRs and the paucity of ligands with specific signaling effects. To better compare the effects of activating different G-protein signaling pathways through ligand-induced or constitutive signaling, we developed a new series of RASSLs (receptors activated solely by synthetic ligands) that activate different G-protein signaling pathways. These RASSLs are based on the human 5-HT(4b) receptor, a GPCR with high constitutive G(s) signaling and strong ligand-induced G-protein activation of the G(s) and G(s/q) pathways. The first receptor in this series, 5-HT(4)-D(100)A or Rs1 (RASSL serotonin 1), is not activated by its endogenous agonist, serotonin, but is selectively activated by the small synthetic molecules GR113808, GR125487, and RO110-0235. All agonists potently induced G(s) signaling, but only a few (e.g., zacopride) also induced signaling via the G(q) pathway. Zacopride-induced G(q) signaling was enhanced by replacing the C-terminus of Rs1 with the C-terminus of the human 5-HT(2C) receptor. Additional point mutations (D(66)A and D(66)N) blocked constitutive G(s) signaling and lowered ligand-induced G(q) signaling. Replacing the third intracellular loop of Rs1 with that of human 5-HT(1A) conferred ligand-mediated G(i) signaling. This G(i)-coupled RASSL, Rs1.3, exhibited no measurable signaling to the G(s) or G(q) pathway. These findings show that the signaling repertoire of Rs1 can be expanded and controlled by receptor engineering and drug selection.

PMID: 18338032 [PubMed - indexed for MEDLINE]

NF-kB signaling pathway NF-kB pathway NF-kB signaling

Altered IkappaBalpha expression promotes NF-kappaB activation in monocytes from primary Sj�gren's syndrome patients.

Altered IkappaBalpha expression promotes NF-kappaB activation in monocytes from primary Sj�gren's syndrome patients.

Pathology. 2012 Aug 29;

Authors: Lisi S, Sisto M, Lofrumento DD, D'Amore M

Abstract
AIMS:: To study the importance of I?B? in NF-?B signal transduction, we analysed the I?B? expression in monocytes from Sj�gren's syndrome (SS) patients versus healthy controls. METHODS:: Monocytes were obtained from the peripheral blood of 30 SS patients and 23 healthy subjects. I?B? expression was studied by semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR), real-time PCR, immunoblotting, flow cytometry and enzyme linked immunosorbent assay (ELISA). RESULTS:: Analysis of the gene and protein expression profiles of SS monocytes revealed a down-regulation of I?B?, and in all the Sj�gren's syndrome cases examined, serum I?B? levels were significantly decreased in comparison with controls. CONCLUSIONS:: Our findings clearly demonstrate changes in the levels of I?B? in SS monocytes, suggesting that the attenuated expression of I?B? could contribute to the deregulation of NF-?B pathways in the SS pathogenesis. Decreased expression of I?B? may specifically amplify cytokines production and inflammatory response linked to Sj�gren's syndrome.

PMID: 22935973 [PubMed - as supplied by publisher]

NF-kB pathway NF-kB signaling NF-kappaB signaling pathway

Subtype-specific roles of phospholipase C-? via differential interactions with PDZ domain proteins.

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Subtype-specific roles of phospholipase C-? via differential interactions with PDZ domain proteins.

Adv Enzyme Regul. 2011;51(1):138-51

Authors: Kim JK, Lim S, Kim J, Kim S, Kim JH, Ryu SH, Suh PG

Abstract
Since we first identified the PLC-? isozyme, enormous studies have been conducted to investigate the functional roles of this protein (Min et al., 1993; Suh et al.,1988). It is now well-known that the four PLC-? subtypes are major effector molecules in GPCR-mediated signaling, especially for intracellular Ca2+ signaling. Nonetheless, it is still poorly understood why multiple PLC-? subtype exist. Most cells express multiple subtypes of PLC-? in different combinations, and each subtype is involved in somewhat different signaling pathways. Therefore, studying the differential roles of each PLC-? subtype is a very interesting issue. In this regard, we focus here on PDZ domain proteins which are novel PLC-? interacting proteins. As scaffolders, PDZ domain proteins recruit various target proteins ranging from membrane receptors to cytoskeletal proteins to assemble highly organized signaling complexes; this can give rise to efficiency and diversity in cellular signaling. Because PLC-? subtypes have different PDZ-binding motifs, it is possible that they are engaged with different PDZ domain proteins, and in turn participate in distinct physiological responses. To date, several PDZ domain proteins, such as the NHERF family, Shank2, and Par-3, have been reported to selectively interact with certain PLC-? subtypes and GPCRs. Systematic predictions of potential binding partners also suggests differential binding properties between PLC-? subtypes. Furthermore, we elucidated parallel signaling processes for multiple PLC-? subtypes, which still perform distinct functions resulting from differential interactions with PDZ domain proteins within a single cell. Therefore, these results highlight the novel function of PDZ domain proteins as intermediaries in subtype-specific role of PLC-? in GPCR-mediated signaling. Future studies will focus on the physiological meanings of this signaling complex formation by different PDZ domain proteins and PLC-? subtypes. It has been observed for a long time that the expression of certain PLC-? subtype fluctuates during diverse physiological conditions. For example, the expression of PLC-?1 is selectively increased during myoblast and adipocyte differentiation (Faenza et al., 2004; O'Carroll et al., 2009). Likewise, PLC-?2 is highly up-regulated during breast cancer progression and plays a critical role in cell migration and mitosis (Bertagnolo et al., 2007). Although PLC-?3 is selectively down-regulated in neuroendocrine tumors, the expression of PLC-?1 is increased in small cell lung carcinoma (Stalberg et al., 2003; Strassheim et al., 2000). In our hypothetical model, it is most likely that up- and down regulation of certain PLC-? subtypes are due to their selective coupling with specific GPCR-mediated signaling, implicated in these pathophysiologic conditions. Therefore, better understanding of selective coupling between PLC-? subtypes, PDZ domain proteins, and GPCRs will shed light on new prognosis and therapy of diverse diseases, and provide potential targets for drug development.

PMID: 21035486 [PubMed - indexed for MEDLINE]

NF-kB signaling NF-kappaB signaling pathway NF-kB signaling pathway

Structure and function analyses of the purified GPCR human vomeronasal type 1 receptor 1.

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Structure and function analyses of the purified GPCR human vomeronasal type 1 receptor 1.

Sci Rep. 2011;1:172

Authors: Corin K, Baaske P, Geissler S, Wienken CJ, Duhr S, Braun D, Zhang S

Abstract
The vomeronasal system is one of several fine-tuned scent-detecting signaling systems in mammals. However, despite significant efforts, how these receptors detect scent remains an enigma. One reason is the lack of sufficient purified receptors to perform detailed biochemical, biophysical and structural analyses. Here we report the ability to express and purify milligrams of purified, functional human vomeronasal receptor hVN1R1. Circular dichroism showed that purified hVN1R1 had an alpha-helical structure, similar to that of other GPCRs. Microscale thermophoresis showed that hVN1R1 bound its known ligand myrtenal with an EC(50) approximately 1 �M. This expression system can enable structural and functional analyses towards understanding how mammalian scent detection works.

PMID: 22355687 [PubMed]

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Proteomics characterization of cytoplasmic and lipid-associated membrane proteins of human pathogen Mycoplasma fermentans M64.

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Proteomics characterization of cytoplasmic and lipid-associated membrane proteins of human pathogen Mycoplasma fermentans M64.

PLoS One. 2012;7(4):e35304

Authors: Liu YC, Lin IH, Chung WJ, Hu WS, Ng WV, Lu CY, Huang TY, Shu HW, Hsiao KJ, Tsai SF, Chang CH, Lin CH

Abstract
Mycoplasma fermentans is a potent human pathogen which has been implicated in several diseases. Notably, its lipid-associated membrane proteins (LAMPs) play a role in immunomodulation and development of infection-associated inflammatory diseases. However, the systematic protein identification of pathogenic M. fermentans has not been reported. From our recent sequencing results of M. fermentans M64 isolated from human respiratory tract, its genome is around 1.1 Mb and encodes 1050 predicted protein-coding genes. In the present study, soluble proteome of M. fermentans was resolved and analyzed using two-dimensional gel electrophoresis. In addition, Triton X-114 extraction was carried out to enrich amphiphilic proteins including putative lipoproteins and membrane proteins. Subsequent mass spectrometric analyses of these proteins had identified a total of 181 M. fermentans ORFs. Further bioinformatics analysis of these ORFs encoding proteins with known or so far unknown orthologues among bacteria revealed that a total of 131 proteins are homologous to known proteins, 11 proteins are conserved hypothetical proteins, and the remaining 39 proteins are likely M. fermentans-specific proteins. Moreover, Triton X-114-enriched fraction was shown to activate NF-kB activity of raw264.7 macrophage and a total of 21 lipoproteins with predicted signal peptide were identified therefrom. Together, our work provides the first proteome reference map of M. fermentans as well as several putative virulence-associated proteins as diagnostic markers or vaccine candidates for further functional study of this human pathogen.

PMID: 22536369 [PubMed - indexed for MEDLINE]

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