Soluble expression in Escherichia coli of active human cyclic nucleotide phosphodiesterase isoform 4B2 in fusion with maltose-binding protein

Recombinant expression in Escherichia coli of human cyclic nucleotide phosphodiesterase 4B2 (hPDE4B2) fused to maltose-binding-protein (MBP-hPDE4B2) was investigated. hPDE4B2 DNA amplified via nested RT-PCR with total RNAs from U937 cells was ligated with pMAL-p2x. After induction at 18 °C for 16 h,...

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Veröffentlicht in:	Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 2009-04, Vol.73 (4), p.968-970
Hauptverfasser:	Zhu, S.(Chongqing Medical Univ. (China)), Yang, G, Yang, X, Zhao, Y, Li, X, Deng, P, Xie, Y, Gan, Z, Liu, Y, Li, Z, Liao, J, Yu, M, Liao, F
Format:	Artikel
Sprache:	eng
Schlagworte:	Animals ANTI-INFLAMMATOIRE ANTIINFLAMMATORY AGENTS ANTINFLAMATORIOS Biological and medical sciences BIOLOGICAL PRODUCTION Carrier Proteins - genetics Cell Extracts Cell Line, Tumor CHEMICOPHYSICAL PROPERTIES cyclic nucleotide phosphodiesterase 4 Cyclic Nucleotide Phosphodiesterases, Type 4 - biosynthesis Cyclic Nucleotide Phosphodiesterases, Type 4 - chemistry Cyclic Nucleotide Phosphodiesterases, Type 4 - genetics Cyclic Nucleotide Phosphodiesterases, Type 4 - isolation & purification ENFERMEDADES RESPIRATORIAS ESCHERICHIA COLI Escherichia coli - genetics EXPRESION GENICA EXPRESSION DES GENES FOSFODIESTERASA Fundamental and applied biological sciences. Psychology GENE EXPRESSION Gene Expression - drug effects GENERO HUMANO GENRE HUMAIN Humans Isoenzymes - biosynthesis Isoenzymes - chemistry Isoenzymes - isolation & purification Isopropyl Thiogalactoside - pharmacology kinetic properties MALADIE RESPIRATOIRE Maltose-Binding Proteins maltose-binding-protein MANKIND PHOSPHODIESTERASE PRODUCCION BIOLOGICA PRODUCTION BIOLOGIQUE PROPIEDADES FISICOQUIMICAS PROPRIETE PHYSICOCHIMIQUE recombinant expression Recombinant Fusion Proteins - biosynthesis Recombinant Fusion Proteins - chemistry Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - isolation & purification RESPIRATORY DISEASES rolipram Solubility
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Zusammenfassung:	Recombinant expression in Escherichia coli of human cyclic nucleotide phosphodiesterase 4B2 (hPDE4B2) fused to maltose-binding-protein (MBP-hPDE4B2) was investigated. hPDE4B2 DNA amplified via nested RT-PCR with total RNAs from U937 cells was ligated with pMAL-p2x. After induction at 18 °C for 16 h, soluble MBP-hPDE4B2 was produced in E. coli. MBP-hPDE4B2 after amylose-resin chromatography showed 35% homogeneity, and its Michaelis-Menten constant was 10±2 μM (n=3). Rolipram had a dissociation constant of 9±2 nM (n=2), and zinc ion was a potent inhibitor. Hence, MBP-hPDE4B2 was expressed in E. coli as a soluble active protein.
ISSN:	0916-8451 1347-6947
DOI:	10.1271/bbb.90131