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Hai-meng Zhou - One of the best experts on this subject based on the ideXlab platform.
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The unfolding intermediate state of calf intestinal Alkaline Phosphatase during denaturation in guanidine solutions
Journal of Protein Chemistry, 2003Co-Authors: Ying-xia Zhang, Xiao-hong Song, Shu-lian Yan, Hai-meng ZhouAbstract:The equilibrium unfolding of calf intestinal Alkaline Phosphatase in guanidinium chloride (GdmCl) solutions was studied by following the fluorescence and ultraviolet difference spectra. At low concentrations of GdmCl (
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Study of refolding of calf intestinal Alkaline Phosphatase.
Journal of Protein Chemistry, 2003Co-Authors: Xiao-juan Tian, Ying-xia Zhang, Xiao-hong Song, Shu-lian Yan, Hai-meng ZhouAbstract:Calf intestinal Alkaline Phosphatase (CIP) was denatured in 3.0 M guanidine hydrochloride for 2 h at 25 degrees C, before being diluted 20-fold with 0.1 M, pH 8.0, Tris-HCl buffer solution containing various effector molecules such as Mg2+, Zn2+, and nucleotide phosphate. The reactivation courses of the enzyme were investigated by the level of activity recovery, the recovery rate constant, and the relative standard deviation of the data. In the presence of effectors, the courses under reducing and nonreducing conditions of disulfide bonds of protein were compared. It was concluded that for CIP, Mg2+ is a more efficient inducer of reconstitution of the active site and appears to play a specific role. In addition, the present study discusses the differences in the refolding effectors between bacterial and mammalian enzymes.
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Effect of extraneous zinc on calf intestinal Alkaline Phosphatase.
Journal of Protein Chemistry, 2003Co-Authors: Shu-lian Yan, Ying-xia Zhang, Xiao-juan Tian, Yong-li Liu, Hai-meng ZhouAbstract:The effect of extraneous zinc on calf intestinal Alkaline Phosphatase was studied for quick reversible binding and slow irreversible binding of zinc ions at various concentrations. Under the conditions of slow binding of zinc to CIP increasing Zn2+ (less than 1.0 mM, nM/nE 1.0 x 10(6)) inhibited enzymatic activity, and further increasing Zn2+ resulted in an increase of activity. For quick reversible binding of Zn2+, the effect on CIP activity changed at lower concentrations of substrate, indicating a complex cooperativity between Zn2+ and pNPP. Both protein intrinsic emission fluorescence and ANS-bound protein fluorescence, as well as circular dichroism spectra have shown that the binding of zinc ions changed the enzyme conformation, which was the reason for the changes in enzyme activity induced by extraneous zinc.
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Comparison of inactivation and unfolding of calf intestinal Alkaline Phosphatase in guanidinium chloride solution
Tsinghua Science and Technology, 2002Co-Authors: Ying-xia Zhang, Hongwei Xi, Hai-meng ZhouAbstract:The changes in activity and unfolding of calf intestinal Alkaline Phosphatase (CIP) during denaturation in guanidinium chloride solutions of different concentrations were investigated using ultraviolet difference absorption spectra and fluorescence emission spectra. Unfolding and inactivation rate constants were measured and compared. The inactivation course is much faster than that of unfolding, which suggests that the active site of CIP containing two zinc ions and one magnesium ion is situated in a limited and flexible region of the enzyme molecule, which is more fragile to the denaturant than the protein as a whole.
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Refolding and reactivation of calf intestinal Alkaline Phosphatase with excess magnesium ions.
The International Journal of Biochemistry & Cell Biology, 2002Co-Authors: Ying-xia Zhang, Yong-li Liu, Ying Zhu, Hai-meng ZhouAbstract:Abstract It is well known that Mg2+ is an essential component in many biological processes. This research investigated the courses of both the reactivation and the refolding in the absence and presence of Mg2+ ions. Calf intestinal Alkaline Phosphatase (CIP) was extensively denatured in 3 M guanidine hydrochloride (GdnHCl) solution for 2 h. Under suitable renaturation conditions, about 60–70% of the activity was recovered in the absence and presence of different magnesium ion concentrations. The refolding processes followed two-phase courses, whereas the reactivation processes were monophasic after dilution in proper solutions with or without Mg2+. The magnesium ions affected both the reactivation and the refolding courses of unfolded CIP. A comparison of rate constants for the refolding of unfolded CIP with those for recovery of enzyme activity at different Mg2+ concentrations showed that they were not synchronized. The activity recovery was speeded up due to the presence of Mg2+ ions; while the refolding course of unfolded CIP was somewhat inhibited by the excess Mg2+.
Ying-xia Zhang - One of the best experts on this subject based on the ideXlab platform.
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Effects of magnesium ions on thermal inactivation of Alkaline Phosphatase.
Protein Journal, 2005Co-Authors: Ying Zhu, Xue-ying Song, Wen-hua Zhao, Ying-xia ZhangAbstract:The effect of Mg2+ on the thermal inactivation and unfolding of calf intestinal Alkaline Phosphatase has been studied at different temperatures and Mg2+ concentrations. Increasing the Mg2+ concentration in the denatured system significantly enhanced the inactivation and unfolding of the enzyme during thermal inactivation. The analysis of the kinetic course of substrate reaction during thermal inactivation showed that at 47°C the increased free Mg2+ concentration caused the inactivation rate to increase. Increasing the temperature strengthened the effect of Mg2+ on the thermal inactivation. Control experiment showed that this is not due to salt effect. The time course of fluorescence emission spectra showed that the emission maximum for Mg2+-containing system was always higher than that of Mg2+-free system, and the higher temperature enhanced this difference. In addition, Mg2+also enhanced the unfolding rate of the enzyme at 47°C. The potential biological significance of these results are discussed.
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Effects of magnesium ions on thermal inactivation of Alkaline Phosphatase.
The Protein Journal, 2005Co-Authors: Ying Zhu, Xue-ying Song, Wen-hua Zhao, Ying-xia ZhangAbstract:The effect of Mg2+ on the thermal inactivation and unfolding of calf intestinal Alkaline Phosphatase has been studied at different temperatures and Mg2+ concentrations. Increasing the Mg2+ concentration in the denatured system significantly enhanced the inactivation and unfolding of the enzyme during thermal inactivation. The analysis of the kinetic course of substrate reaction during thermal inactivation showed that at 47 degrees C the increased free Mg2+ concentration caused the inactivation rate to increase. Increasing the temperature strengthened the effect of Mg2+ on the thermal inactivation. Control experiment showed that this is not due to salt effect. The time course of fluorescence emission spectra showed that the emission maximum for Mg2+-containing system was always higher than that of Mg2+-free system, and the higher temperature enhanced this difference. In addition, Mg2+ also enhanced the unfolding rate of the enzyme at 47 degrees C. The potential biological significance of these results are discussed.
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The unfolding intermediate state of calf intestinal Alkaline Phosphatase during denaturation in guanidine solutions
Journal of Protein Chemistry, 2003Co-Authors: Ying-xia Zhang, Xiao-hong Song, Shu-lian Yan, Hai-meng ZhouAbstract:The equilibrium unfolding of calf intestinal Alkaline Phosphatase in guanidinium chloride (GdmCl) solutions was studied by following the fluorescence and ultraviolet difference spectra. At low concentrations of GdmCl (
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Study of refolding of calf intestinal Alkaline Phosphatase.
Journal of Protein Chemistry, 2003Co-Authors: Xiao-juan Tian, Ying-xia Zhang, Xiao-hong Song, Shu-lian Yan, Hai-meng ZhouAbstract:Calf intestinal Alkaline Phosphatase (CIP) was denatured in 3.0 M guanidine hydrochloride for 2 h at 25 degrees C, before being diluted 20-fold with 0.1 M, pH 8.0, Tris-HCl buffer solution containing various effector molecules such as Mg2+, Zn2+, and nucleotide phosphate. The reactivation courses of the enzyme were investigated by the level of activity recovery, the recovery rate constant, and the relative standard deviation of the data. In the presence of effectors, the courses under reducing and nonreducing conditions of disulfide bonds of protein were compared. It was concluded that for CIP, Mg2+ is a more efficient inducer of reconstitution of the active site and appears to play a specific role. In addition, the present study discusses the differences in the refolding effectors between bacterial and mammalian enzymes.
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Effect of extraneous zinc on calf intestinal Alkaline Phosphatase.
Journal of Protein Chemistry, 2003Co-Authors: Shu-lian Yan, Ying-xia Zhang, Xiao-juan Tian, Yong-li Liu, Hai-meng ZhouAbstract:The effect of extraneous zinc on calf intestinal Alkaline Phosphatase was studied for quick reversible binding and slow irreversible binding of zinc ions at various concentrations. Under the conditions of slow binding of zinc to CIP increasing Zn2+ (less than 1.0 mM, nM/nE 1.0 x 10(6)) inhibited enzymatic activity, and further increasing Zn2+ resulted in an increase of activity. For quick reversible binding of Zn2+, the effect on CIP activity changed at lower concentrations of substrate, indicating a complex cooperativity between Zn2+ and pNPP. Both protein intrinsic emission fluorescence and ANS-bound protein fluorescence, as well as circular dichroism spectra have shown that the binding of zinc ions changed the enzyme conformation, which was the reason for the changes in enzyme activity induced by extraneous zinc.
Mubashir Hassan - One of the best experts on this subject based on the ideXlab platform.
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Understanding the enzymatic inhibition of intestinal Alkaline Phosphatase by aminophenazone-derived aryl thioureas with aided computational molecular dynamics simulations: synthesis, characterization, SAR and kinetic profiling
Molecular Diversity, 2020Co-Authors: Asma Khurshid, Aamer Saeed, Qamar Abbas, Zaman Ashraf, Mubashir HassanAbstract:The work presented in this paper aims toward the synthesis of aryl thiourea derivatives 4a – l of pyrazole based nonsteroidal anti-inflammatory drug named 4-aminophenazone, as potential inhibitors of intestinal Alkaline Phosphatase enzyme. The screening of synthesized target compounds 4a – l for unraveling the anti-inflammatory potential against calf intestinal Alkaline Phosphatase gives rise to lead member 4c possessing IC_50 value 0.420 ± 0.012 µM, many folds better than reference standard used (KH_2PO_4 IC_50 = 2.8 ± 0.06 µM and l -phenylalanine IC_50 = 100 ± 3.1 µM). SAR for unfolding the active site binding pocket interaction along with the mode of enzyme inhibition based on kinetic studies is carried out which showed non-competitive binding mode. The enzyme inhibition studies were further supplemented by molecular dynamic simulations for predicting the protein behavior against active inhibitors 4c and 4g during docking analysis. The preliminary toxicity of the synthesized compounds was determined by using brine shrimp assay. This work also includes detailed biochemical analysis along with RO5 parameters for all the newly synthesized drug derivatives 4a – l . Graphic abstract
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Synthesis of sulfadiazinyl acyl/aryl thiourea derivatives as calf intestinal Alkaline Phosphatase inhibitors, pharmacokinetic properties, lead optimization, Lineweaver-Burk plot evaluation and binding analysis.
Bioorganic & Medicinal Chemistry, 2018Co-Authors: Sajid-ur-rehman, Aamer Saeed, Gufran Saddique, Pervaiz Ali Channar, Fayaz Ali Larik, Qamar Abbas, Mubashir Hassan, Hussain Raza, Tanzeela Abdul Fattah, Sung-yum SeoAbstract:Abstract To seek the new medicinal potential of sulfadiazine drug, the free amino group of sulfadiazine was exploited to obtain acyl/aryl thioureas using simple and straightforward protocol. Acyl/aryl thioureas are well recognized bioactive pharmacophore containing moieties. A new series (4a–4j) of sulfadiazine derived acyl/aryl thioureas was synthesized and characterized through spectroscopic and elemental analysis. The synthesized derivatives 4a–4j were subjected to calf intestinal Alkaline Phosphatase (CIAP) activity. The derivative 4a–4j showed better inhibition potential compared to standard monopotassium phosphate (MKP). The compound 4c exhibited higher potential in the series with IC50 0.251 ± 0.012 µM (standard KH2PO4 4.317 ± 0.201 µM). Lineweaver-Burk plots revealed that most potent derivative 4c inhibition CIAP via mixed type pathway. Pharmacological investigations showed that synthesized compounds 4a–4j obey Lipinsk’s rule. ADMET parameters evaluation predicted that these molecule show significant lead like properties with minimum possible toxicity and can serve as templates in drug designing. The synthetic compounds show none mutagenic and irritant behavior. Molecular docking analysis showed that compound 4c interacts with Asp273, His317 and Arg166 amino acid residues.
Qamar Abbas - One of the best experts on this subject based on the ideXlab platform.
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Understanding the enzymatic inhibition of intestinal Alkaline Phosphatase by aminophenazone-derived aryl thioureas with aided computational molecular dynamics simulations: synthesis, characterization, SAR and kinetic profiling
Molecular Diversity, 2020Co-Authors: Asma Khurshid, Aamer Saeed, Qamar Abbas, Zaman Ashraf, Mubashir HassanAbstract:The work presented in this paper aims toward the synthesis of aryl thiourea derivatives 4a – l of pyrazole based nonsteroidal anti-inflammatory drug named 4-aminophenazone, as potential inhibitors of intestinal Alkaline Phosphatase enzyme. The screening of synthesized target compounds 4a – l for unraveling the anti-inflammatory potential against calf intestinal Alkaline Phosphatase gives rise to lead member 4c possessing IC_50 value 0.420 ± 0.012 µM, many folds better than reference standard used (KH_2PO_4 IC_50 = 2.8 ± 0.06 µM and l -phenylalanine IC_50 = 100 ± 3.1 µM). SAR for unfolding the active site binding pocket interaction along with the mode of enzyme inhibition based on kinetic studies is carried out which showed non-competitive binding mode. The enzyme inhibition studies were further supplemented by molecular dynamic simulations for predicting the protein behavior against active inhibitors 4c and 4g during docking analysis. The preliminary toxicity of the synthesized compounds was determined by using brine shrimp assay. This work also includes detailed biochemical analysis along with RO5 parameters for all the newly synthesized drug derivatives 4a – l . Graphic abstract
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Synthesis of sulfadiazinyl acyl/aryl thiourea derivatives as calf intestinal Alkaline Phosphatase inhibitors, pharmacokinetic properties, lead optimization, Lineweaver-Burk plot evaluation and binding analysis.
Bioorganic & Medicinal Chemistry, 2018Co-Authors: Sajid-ur-rehman, Aamer Saeed, Gufran Saddique, Pervaiz Ali Channar, Fayaz Ali Larik, Qamar Abbas, Mubashir Hassan, Hussain Raza, Tanzeela Abdul Fattah, Sung-yum SeoAbstract:Abstract To seek the new medicinal potential of sulfadiazine drug, the free amino group of sulfadiazine was exploited to obtain acyl/aryl thioureas using simple and straightforward protocol. Acyl/aryl thioureas are well recognized bioactive pharmacophore containing moieties. A new series (4a–4j) of sulfadiazine derived acyl/aryl thioureas was synthesized and characterized through spectroscopic and elemental analysis. The synthesized derivatives 4a–4j were subjected to calf intestinal Alkaline Phosphatase (CIAP) activity. The derivative 4a–4j showed better inhibition potential compared to standard monopotassium phosphate (MKP). The compound 4c exhibited higher potential in the series with IC50 0.251 ± 0.012 µM (standard KH2PO4 4.317 ± 0.201 µM). Lineweaver-Burk plots revealed that most potent derivative 4c inhibition CIAP via mixed type pathway. Pharmacological investigations showed that synthesized compounds 4a–4j obey Lipinsk’s rule. ADMET parameters evaluation predicted that these molecule show significant lead like properties with minimum possible toxicity and can serve as templates in drug designing. The synthetic compounds show none mutagenic and irritant behavior. Molecular docking analysis showed that compound 4c interacts with Asp273, His317 and Arg166 amino acid residues.
Aamer Saeed - One of the best experts on this subject based on the ideXlab platform.
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Understanding the enzymatic inhibition of intestinal Alkaline Phosphatase by aminophenazone-derived aryl thioureas with aided computational molecular dynamics simulations: synthesis, characterization, SAR and kinetic profiling
Molecular Diversity, 2020Co-Authors: Asma Khurshid, Aamer Saeed, Qamar Abbas, Zaman Ashraf, Mubashir HassanAbstract:The work presented in this paper aims toward the synthesis of aryl thiourea derivatives 4a – l of pyrazole based nonsteroidal anti-inflammatory drug named 4-aminophenazone, as potential inhibitors of intestinal Alkaline Phosphatase enzyme. The screening of synthesized target compounds 4a – l for unraveling the anti-inflammatory potential against calf intestinal Alkaline Phosphatase gives rise to lead member 4c possessing IC_50 value 0.420 ± 0.012 µM, many folds better than reference standard used (KH_2PO_4 IC_50 = 2.8 ± 0.06 µM and l -phenylalanine IC_50 = 100 ± 3.1 µM). SAR for unfolding the active site binding pocket interaction along with the mode of enzyme inhibition based on kinetic studies is carried out which showed non-competitive binding mode. The enzyme inhibition studies were further supplemented by molecular dynamic simulations for predicting the protein behavior against active inhibitors 4c and 4g during docking analysis. The preliminary toxicity of the synthesized compounds was determined by using brine shrimp assay. This work also includes detailed biochemical analysis along with RO5 parameters for all the newly synthesized drug derivatives 4a – l . Graphic abstract
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Synthesis of sulfadiazinyl acyl/aryl thiourea derivatives as calf intestinal Alkaline Phosphatase inhibitors, pharmacokinetic properties, lead optimization, Lineweaver-Burk plot evaluation and binding analysis.
Bioorganic & Medicinal Chemistry, 2018Co-Authors: Sajid-ur-rehman, Aamer Saeed, Gufran Saddique, Pervaiz Ali Channar, Fayaz Ali Larik, Qamar Abbas, Mubashir Hassan, Hussain Raza, Tanzeela Abdul Fattah, Sung-yum SeoAbstract:Abstract To seek the new medicinal potential of sulfadiazine drug, the free amino group of sulfadiazine was exploited to obtain acyl/aryl thioureas using simple and straightforward protocol. Acyl/aryl thioureas are well recognized bioactive pharmacophore containing moieties. A new series (4a–4j) of sulfadiazine derived acyl/aryl thioureas was synthesized and characterized through spectroscopic and elemental analysis. The synthesized derivatives 4a–4j were subjected to calf intestinal Alkaline Phosphatase (CIAP) activity. The derivative 4a–4j showed better inhibition potential compared to standard monopotassium phosphate (MKP). The compound 4c exhibited higher potential in the series with IC50 0.251 ± 0.012 µM (standard KH2PO4 4.317 ± 0.201 µM). Lineweaver-Burk plots revealed that most potent derivative 4c inhibition CIAP via mixed type pathway. Pharmacological investigations showed that synthesized compounds 4a–4j obey Lipinsk’s rule. ADMET parameters evaluation predicted that these molecule show significant lead like properties with minimum possible toxicity and can serve as templates in drug designing. The synthetic compounds show none mutagenic and irritant behavior. Molecular docking analysis showed that compound 4c interacts with Asp273, His317 and Arg166 amino acid residues.
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Isonicotinohydrazones as inhibitors of Alkaline Phosphatase and ecto-5′-nucleotidase
Chemical Biology & Drug Design, 2016Co-Authors: Pervaiz Ali Channar, Aamer Saeed, Syed Jawad Ali Shah, Sidra Hassan, Zaib Un Nisa, Joanna Lecka, Jean Sévigny, Jürgen Bajorath, Jamshed IqbalAbstract:A series of isonicotinohydrazide derivatives was synthesized and tested against recombinant human and rat ecto-5′-nucleotidases (h-e5′NT and r-e5′NT) and Alkaline Phosphatase isozymes including both bovine tissue-non-specific Alkaline Phosphatase (b-TNAP) and tissue-specific calf intestinal Alkaline Phosphatase (c-IAP). These enzymes are implicated in vascular calcifications, hypophosphatasia, solid tumors, and cancers, such as colon, lung, breast, pancreas, and ovary. All tested compounds were active against both enzymes. The most potent inhibitor of h-e5′NT was derivative (E)-N′-(1-(3-(4-fluorophenyl)-5-phenyl-4,5-dihydro-1H-pyrazol-1-yl)ethylidene)isonicotinohydrazide (3j), whereas derivative (E)-N′-(4-hydroxy-3-methoxybenzylidene)isonicotinohydrazide (3g) exhibited significant inhibitory activity against r-e5′NT. In addition, the derivative (E)-N′-(4′-chlorobenzylidene)isonicotinohydrazide (3a) was most potent inhibitor against calf intestinal Alkaline Phosphatase and the derivative (E)-N′-(4-hydroxy-3-methoxybenzylidene)isonicotinohydrazide (3g) was found to be most potent inhibitor of bovine tissue-non-specific Alkaline Phosphatase. Furthermore, putative binding modes of potent compounds against e5′NT (human and rat e5′NT) and AP (including b-TNAP and c-IAP) were determined computationally.
