A synthetic glucagon-like peptide-1 analog with improved plasma stability

Glucagon-like peptide-1 (GLP-1) is the most potent endogenous insulin-stimulating hormone. In the present study the plasma stability and biological activity of a GLP-1 analog, [Ser]GLP-1(7-36)amide, in which the second N-terminal amino acid alanine was replaced by serine, was evaluated in vitro and in vivo. Incubation of GLP-1 with human or rat plasma resulted in degradation of native GLP-1(7-36)amide to GLP-1(9-36)amide, while [Ser]GLP-1(7-36)amide was not significantly degraded by plasma enzymes. Using glucose-responsive HIT-T15 cells, [Ser]GLP-1(7-36)amide showed strong insulinotropic activity, which was inhibited by the specific GLP-1 receptor antagonist exendin-4(9-39)amide. Simultaneous i.v. injection of [Ser]GLP-1(7-36)amide and glucose in rats induced a twofold higher increase in plasma insulin levels than unmodified GLP-1(7-36)amide with glucose and a fivefold higher increase than glucose alone. [Ser]GLP-1(7-36)amide induced a 1.5-fold higher increase in plasma insulin than GLP-1(7-36)amide when given 1 h before i.v. application of glucose. The insulinotropic effect of [Ser]GLP-1(7-36)amide was suppressed by i.v. application of exendin-4(9-39)amide. The present data demonstrate that replacement of the second N-terminal amino acid alanine by serine improves the plasma stability of GLP-1(7-36)amide. The insulinotropic action in vitro and in vivo was not impaired significantly by this modification.

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Effects of substitution of n-terminal amino acid of glucagon-like peptide-1 (7-36) amide on food intake of the neonatal chick

Life Sciences ◽

10.1016/s0024-3205(99)00535-4 ◽

1999 ◽

Vol 65 (24) ◽

pp. PL295-PL299

Author(s):

Takashi Bungo ◽

Masataka Shimojo ◽

Yasuhisa Masuda ◽

Noboru Saito ◽

Kunio Sugahara ◽

...

Keyword(s):

Amino Acid ◽

Food Intake ◽

Terminal Amino Acid ◽

Neonatal Chick ◽

Terminal Amino ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

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Energy-dependent protein folding: modeling how a protein folding machine may work

F1000Research ◽

10.12688/f1000research.28175.1 ◽

2021 ◽

Vol 10 ◽

pp. 3

Author(s):

Harutyun Sahakian ◽

Karen Nazarian ◽

Arcady Mushegian ◽

Irina Sorokina

Keyword(s):

Molecular Dynamics ◽

Protein Folding ◽

Amino Acid ◽

Dynamics Simulation ◽

Terminal Amino Acid ◽

Peptide Backbone ◽

Terminal Amino ◽

Energy Dependent

Background: Proteins fold robustly and reproducibly in vivo, but many cannot fold in vitro in isolation from cellular components. Despite the remarkable progress that has been achieved by the artificial intelligence approaches in predicting the protein native conformations, the pathways that lead to such conformations, either in vitro or in vivo, remain largely unknown. The slow progress in recapitulating protein folding pathways in silico may be an indication of the fundamental deficiencies in our understanding of folding as it occurs in nature. Here we consider the possibility that protein folding in living cells may not be driven solely by the decrease in Gibbs free energy and propose that protein folding in vivo should be modeled as an active energy-dependent process. The mechanism of action of such a protein folding machine might include direct manipulation of the peptide backbone. Methods: To show the feasibility of a protein folding machine, we conducted molecular dynamics simulations that were augmented by the application of mechanical force to rotate the C-terminal amino acid while simultaneously limiting the N-terminal amino acid movements. Results: Remarkably, the addition of this simple manipulation of peptide backbones to the standard molecular dynamics simulation indeed facilitated the formation of native structures in five diverse alpha-helical peptides. Steric clashes that arise in the peptides due to the forced directional rotation resulted in the behavior of the peptide backbone no longer resembling a freely jointed chain. Conclusions: These simulations show the feasibility of a protein folding machine operating under the conditions when the movements of the polypeptide backbone are restricted by applying external forces and constraints. Further investigation is needed to see whether such an effect may play a role during co-translational protein folding in vivo and how it can be utilized to facilitate folding of proteins in artificial environments.

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Energy-dependent protein folding: modeling how a protein folding machine may work

10.1101/2020.09.01.277582 ◽

2020 ◽

Author(s):

Harutyun K. Sahakyan ◽

Karen B. Nazaryan ◽

Arcady R. Mushegian ◽

Irina N. Sorokina

Keyword(s):

Molecular Dynamics ◽

Protein Folding ◽

Amino Acid ◽

Dynamics Simulation ◽

Terminal Amino Acid ◽

Peptide Backbone ◽

Terminal Amino ◽

Energy Dependent

AbstractProteins fold robustly and reproducibly in vivo, but many cannot fold in vitro in isolation from cellular components. The pathways to proteins’ native conformations, either in vitro or in vivo, remain largely unknown. The slow progress in recapitulating protein folding pathways in silico may be an indication of the fundamental deficiencies in our understanding of folding as it occurs in nature. Here we consider the possibility that protein folding in living cells may not be driven solely by the decrease in Gibbs free energy and propose that protein folding in vivo should be modeled as an active energy-dependent process. The mechanism of action of such protein folding machine might include direct manipulation of the peptide backbone. To show the feasibility of a protein folding machine, we conducted molecular dynamics simulations that were augmented by the application of mechanical force to rotate the C-terminal amino acid while simultaneously limiting the N-terminal amino acid movements. Remarkably, the introduction of this simple manipulation of peptide backbones to the standard molecular dynamics simulation indeed facilitated the formation of native structures in five diverse alpha-helical peptides. Such effect may play a role during co-translational protein folding in vivo: considering the rotating motion of the tRNA 3’-end in the peptidyltransferase center of the ribosome, it is possible that this motion might introduce rotation to the nascent peptide and influence the peptide’s folding pathway in a way similar to what was observed in our simulations.

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The major kidney AE1 isoform does not bind ankyrin (Ank1) in vitro. An essential role for the 79 NH2-terminal amino acid residues of band 3.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(18)31621-1 ◽

1994 ◽

Vol 269 (51) ◽

pp. 32201-32208

Author(s):

Y. Ding ◽

J.R. Casey ◽

R.R. Kopito

Keyword(s):

Amino Acid ◽

Essential Role ◽

Band 3 ◽

Amino Acid Residues ◽

Terminal Amino Acid ◽

Terminal Amino

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Glucagon-like Peptide 1 Receptor Agonists, Diabetic Retinopathy and Angiogenesis: The AngioSafe Type 2 Diabetes Study

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/clinem/dgz069 ◽

2019 ◽

Vol 105 (4) ◽

pp. e1549-e1560 ◽

Cited By ~ 7

Author(s):

Bénédicte Gaborit ◽

Jean-Baptiste Julla ◽

Samaher Besbes ◽

Matthieu Proust ◽

Clara Vincentelli ◽

...

Keyword(s):

Type 2 Diabetes ◽

Diabetic Retinopathy ◽

Fundus Photography ◽

Endothelial Cell Proliferation ◽

Receptor Agonists ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Abstract Aims Recent trials provide conflicting results on the association between glucagon-like peptide 1 receptor agonists (GLP-1RA) and diabetic retinopathy (DR). The aim of the AngioSafe type 2 diabetes (T2D) study was to determine the role of GLP-1RA in angiogenesis using clinical and preclinical models. Methods We performed two studies in humans. In study 1, we investigated the effect of GLP-1RA exposure from T2D diagnosis on the severity of DR, as diagnosed with retinal imaging (fundus photography). In study 2, a randomized 4-week trial, we assessed the effect of liraglutide on circulating hematopoietic progenitor cells (HPCs), and angio-miRNAs. We then studied the experimental effect of Exendin-4, on key steps of angiogenesis: in vitro on human endothelial cell proliferation, survival and three-dimensional vascular morphogenesis; and in vivo on ischemia-induced neovascularization of the retina in mice. Results In the cohort of 3154 T2D patients, 10% displayed severe DR. In multivariate analysis, sex, disease duration, glycated hemoglobin (HbA1c), micro- and macroangiopathy, insulin therapy and hypertension remained strongly associated with severe DR, while no association was found with GLP-1RA exposure (o 1.139 [0.800–1.622], P = .47). We further showed no effect of liraglutide on HPCs, and angio-miRNAs. In vitro, we demonstrated that exendin-4 had no effect on proliferation and survival of human endothelial cells, no effect on total length and number of capillaries. Finally, in vivo, we showed that exendin-4 did not exert any negative effect on retinal neovascularization. Conclusions The AngioSafe T2D studies provide experimental and clinical data confirming no effect of GLP-1RA on angiogenesis and no association between GLP-1 exposure and severe DR.

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Oral intake of slowly digestible α-glucan, isomaltodextrin, stimulates glucagon-like peptide-1 secretion in the small intestine of rats

British Journal Of Nutrition ◽

10.1017/s0007114519003210 ◽

2019 ◽

Vol 123 (6) ◽

pp. 619-626

Author(s):

Yoshihiko Komuro ◽

Takashi Kondo ◽

Shingo Hino ◽

Tatsuya Morita ◽

Naomichi Nishimura

Keyword(s):

Small Intestine ◽

Oral Delivery ◽

Oral Intake ◽

Membrane Vesicles ◽

Rat Small Intestine ◽

Maize Starch ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

AbstractTo investigate whether oral intake of highly branched α-glucan isomaltodextrin (IMD) could stimulate ileal glucagon-like peptide-1 (GLP-1) secretion, we examined (1) the digestibility of IMD, (2) the digestion and absorption rates of IMD, in rat small intestine and (3) portal GLP-1 concentration in rats given IMD. In Expt 1, ileorectostomised rats were given a 3 % IMD diet for 10 d. Separately, a 16-h in vitro digestion of IMD, using porcine pancreatic α-amylase and brush-border membrane vesicles from rat small intestine, was conducted. In Expt 2, upon 24-h fasting, rats were given any of glucose, IMD and high-amylose maize starch (HAMS) (1 g/kg of body weight). In Expt 3, caecectomised rats were given 0·2 % neomycin sulphate and a 5 % IMD diet for 10 d. The in vivo and in vitro digestibility of IMD was 70–80 %. The fraction of IMD digested in vitro for the first 120 min was 67 % of that in maize starch. The AUC for 0–120 min of plasma glucose concentration was significantly lower in HAMS group and tended to be lower in IMD group than in the glucose group. Finally, we also observed that, when compared with control rats, glucose of IMD significantly stimulated and improved the concentration of portal active GLP-1 in antibiotic-administered, caecectomised rats. We concluded that IMD was slowly digested and the resulting glucose stimulated GLP-1 secretion in rat small intestine. Oral delivery of slowly released IMD glucose to the small intestine probably exerts important, yet unknown, physiological effects on the recipient.

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Activation of glucagon-like peptide-1 receptor inhibits growth and promotes apoptosis of human pancreatic cancer cells in a cAMP-dependent manner

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00017.2014 ◽

2014 ◽

Vol 306 (12) ◽

pp. E1431-E1441 ◽

Cited By ~ 19

Author(s):

Hejun Zhao ◽

Rui Wei ◽

Liang Wang ◽

Qing Tian ◽

Ming Tao ◽

...

Keyword(s):

Pancreatic Cancer ◽

Cancer Cells ◽

Human Pancreatic Cancer ◽

Dependent Manner ◽

Pancreatic Cancer Cells ◽

Tumor Tissues ◽

Glucagon Like Peptide ◽

Glucagon Like Peptide 1

Glucagon-like peptide-1 (GLP-1) promotes pancreatic β-cell regeneration through GLP-1 receptor (GLP-1R) activation. However, whether it promotes exocrine pancreas growth and thereby increases the risk of pancreatic cancer has been a topic of debate in recent years. Clinical data and animal studies published so far have been controversial. In the present study, we report that GLP-1R activation with liraglutide inhibited growth and promoted apoptosis in human pancreatic cancer cell lines in vitro and attenuated pancreatic tumor growth in a mouse xenograft model in vivo. These effects of liraglutide were mediated through activation of cAMP production and consequent inhibition of Akt and ERK1/2 signaling pathways in a GLP-1R-dependent manner. Moreover, we examined GLP-1R expression in human pancreatic cancer tissues and found that 43.3% of tumor tissues were GLP-1R-null. In the GLP-1R-positive tumor tissues (56.7%), the level of GLP-1R was lower compared with that in tumor-adjacent normal pancreatic tissues. Furthermore, the GLP-1R-positive tumors were significantly smaller than the GLP-1R-null tumors. Our study shows for the first time that GLP-1R activation has a cytoreductive effect on human pancreatic cancer cells in vitro and in vivo, which may help address safety concerns of GLP-1-based therapies in the context of human pancreatic cancer.

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