Poly(ethylene glycol)-Mediated Conformational Alteration of α-Chymotrypsin Prevents Inactivation of Insulin by Stabilizing Active Intermediates

Yu, Jibing; Wei, Xiuli; Zhang, Li; Fang, Xiaocui; Yang, Tao; Huang, Feng; Liang, Wei

doi:10.1021/mp500001n

Cited by 2 publications

(2 citation statements)

References 44 publications

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“…A similar effect was observed for cholesterol oxidases (ChOx), which adsorbed onto PS/PEG particles but presented no enzymatic activity [18]. In solution the effect of PEG on the enzyme conformation and activity seems to depend on the enzyme type and on PEG molecular weight, for instance, PEG 400 changed the conformation of chymotrypsin, retarding the degradation of insulin [32], the enzymatic activity of hexokinase increased 6% in the presence of PEG 400 and decreased in the range 14%–7% in the presence of PEG 1500 and PEG 4000, respectively, due to aggregation [33], PEG 400 or 4000 presented no interaction with glucose-6-phosphate dehydrogenase (G-6-PDH), but favorable interactions were observed between PEG and the co-enzyme NADP + , which increased the enzymatic activity in 20% [34]. Comparing the activities in the presence of PEG 200, PEG 8000 or PEG 12000, the enzymatic activity of lipase increased 161% in the presence of PEG 12000 [35].…”

Section: Resultsmentioning

confidence: 78%

Catalytic Behavior of Lipase Immobilized onto Congo Red and PEG-Decorated Particles

2014

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Poly(ethylene glycol) (PEG)-decorated polystyrene (PS) nanoparticles with mean hydrodynamic diameter (D) and zeta-potential (ζ) of (286 ± 15) nm and (−50 ± 5) mV, respectively, were modified by the adsorption of Congo red (CR). The PS/PEG/CR particles presented D and ζ values of (290 ± 19) nm and (−36 ± 5) mV, respectively. The adsorption of lipase onto PS/PEG or PS/PEG/CR particles at (24 ± 1) °C and pH 7 changed the mean D value to (380 ± 20) and (405 ± 11) nm, respectively, and ζ value to (−32 ± 4) mV and (−25 ± 2) mV, respectively. The kinetic parameters of the hydrolysis of p-nitrophenyl butyrate were determined for free lipase, lipase immobilized onto PS/PEG and PS/PEG/CR particles. Lipase on PS/PEG/CR presented the largest Michaelis-Menten constant (K M ), but also the highest V max and k cat values. Moreover, it could be recycled seven times, losing a maximum 10% or 30% of the original enzymatic activity at 40 °C or 25 °C, respectively. Although lipases immobilized onto PS/PEG particles presented the smallest K M values, the reactions were comparatively the slowest and recycling was not possible. Hydrolysis reactions performed in the temperature range of 25 °C to 60 °C with free lipases and lipases immobilized onto PS/PEG/CR particles presented an optimal temperature at 40 °C. At 60 °C free lipases and lipases immobilized onto PS/PEG/CR presented ~80% and ~50% of the activity measured at 40 °C, indicating good thermal stability. Bioconjugation effects between CR and lipase were evidenced by circular dichroism spectroscopy and spectrophotometry. CR molecules mediate the open state conformation of the lipase lid and favor the substrate approaching.

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Section: Resultsmentioning

confidence: 78%

Catalytic Behavior of Lipase Immobilized onto Congo Red and PEG-Decorated Particles

2014

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“…Inactivation issues can be solved by the use of genetically engineered food-grade microorganisms that secrete therapeutic protein drugs (Yu et al, 2014) or by microencapsulation of the therapeutic protein drug (Shin and Yoo, 2013) to target the therapeutic proteins to the absorption site. However, because of the general large molecular size and hydrophilicity of therapeutic proteins, the issue of malabsorption is difficult to solve.…”

Section: Discussionmentioning

confidence: 99%

Insulin receptor binding motif tagged with IgG4 Fc (Yiminsu) works as an insulin sensitizer to activate Akt signaling in hepatocytes

Wang¹,

T²,

Yang³

et al. 2015

Genet. Mol. Res.

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ABSTRACT. Insulin resistance is a key feature of obesity and type 2 diabetes mellitus (T2DM). Interaction of insulin with the insulin receptor (IR) leads to both its auto-phosphorylation and phosphorylation of tyrosine residues on the IR substrate (IRS) proteins, initiating the activation of intracellular signaling cascades. The metabolic effects of IRS are known to be mediated through pathways involving phosphatidyl-J. Wang et al. 8820©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 14 (3): 8819-8828 (2015) inositol 3-kinase (PI-3K), which result in the activation of Akt signaling. The C-terminal region of the IR ectodomain is required to facilitate the conformational changes that are required for high-affinity binding to insulin. Furthermore, the CH2 and CH3 domains in the Fc fragments of immunoglobulins are responsible for their binding to the Fc receptor, which triggers transcytosis. In this study, we created a fusion peptide of the C-terminal end of the human IR ectodomain with the IgG4 Fc fragment, including an intervening polyG fragment to ensure enough space for insulin binding. We named this new peptide "Yiminsu", meaning an insulin sensitizer. The results of our analyses show that Yiminsu significantly facilitates insulin signaling via the activation of Akt in hepatocytes in a dose-and time-dependent manner. Further studies are required to determine whether Yiminsu can act as an insulin sensitizer.

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Poly(ethylene glycol)-Mediated Conformational Alteration of α-Chymotrypsin Prevents Inactivation of Insulin by Stabilizing Active Intermediates

Cited by 2 publications

References 44 publications

Catalytic Behavior of Lipase Immobilized onto Congo Red and PEG-Decorated Particles

Catalytic Behavior of Lipase Immobilized onto Congo Red and PEG-Decorated Particles

Insulin receptor binding motif tagged with IgG4 Fc (Yiminsu) works as an insulin sensitizer to activate Akt signaling in hepatocytes

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