Cloning, Purification, and Partial Characterization of Bacillus subtilis Urate Oxidase Expressed in Escherichia coli

被引：34

作者：

Pfrimer, Pollyana ^{[1
]}

Pepe de Moraes, Lidia Maria ^{[1
]}

Galdino, Alexsandro Sobreira ^{[1
]}

Salles, Loise Pedrosa ^{[1
]}

Branco Reis, Viviane Castelo ^{[1
]}

De Marco, Janice Lisboa ^{[1
]}

Prates, Maura Vianna ^{[2
]}

Bloch, Carlos, Jr. ^{[2
]}

Goncalves Torres, Fernando Araripe ^{[1
]}

机构：

[1] Univ Brasilia, Mol Biol Lab, BR-70910900 Brasilia, DF, Brazil

[2] EMBRAPA Recursos Genet & Biotecnol, Lab Espectrometria Massa, BR-70770917 Brasilia, DF, Brazil

来源：

JOURNAL OF BIOMEDICINE AND BIOTECHNOLOGY | 2010年

关键词：

SERUM URIC-ACID; EVOLUTIONARY IMPLICATIONS; HYPERURICEMIA; RASBURICASE; LEUKEMIA; SEQUENCE; PRODUCT; GENE; GOUT;

D O I：

10.1155/2010/674908

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Urate oxidase (EC 1.7.3.3) is an enzyme involved in purine metabolism which is used in the treatment of gout and as diagnostic reagent for detection of uric acid. In order to produce this enzyme in large quantities for biotechnological purposes, the gene coding for the Bacillus subtilis urate oxidase was cloned and heterologously expressed in Escherichia coli. Time course induction in E. coli showed an induced protein with an apparent molecular mass of similar to 60 kDa. Soluble recombinant enzyme was purified in a single-step procedure using Ni-NTA column. The enzyme was purified 2.1-fold with a yield of 56% compared to the crude extract. MALDI-TOF analysis revealed an ion with a mass of 58675 Da which is in agreement with the expected mass of the recombinant protein. The purified enzyme showed an optimal pH and temperature of 8.0 and 37 degrees C, respectively, and retained 90% of its activity after 72 hours of incubation at similar to 20 degrees C and 4 degrees C.

引用

页数：6

共 31 条

[1] Hypouricemia and hyperuricemia in type 2 diabetes: two different phenotypes [J].

Bo, S ;

Cavallo-Perin, P ;

Gentile, L ;

Repetti, E ;

Pagano, G .

EUROPEAN JOURNAL OF CLINICAL INVESTIGATION, 2001, 31 (04) :318-321

[2] URIC-ACID DEGRADATION BY BACILLUS-FASTIDIOSUS STRAINS [J].

BONGAERTS, GPA ;

VOGELS, GD .

JOURNAL OF BACTERIOLOGY, 1976, 125 (02) :689-697

[3]

BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3

[4]

Cammalleri Lisa, 2007, Int J Med Sci, V4, P83

[5] Update on emerging urate-lowering therapies [J].

Chohan, Saima ;

Becker, Michael A. .

CURRENT OPINION IN RHEUMATOLOGY, 2009, 21 (02) :143-149

[6] A colorimetric 96-well microtiter plate assay for the determination of urate oxidase activity and its kinetic parameters [J].

Fraisse, L ;

Bonnet, MC ;

de Farcy, JP ;

Agut, C ;

Dersigny, D ;

Bayol, A .

ANALYTICAL BIOCHEMISTRY, 2002, 309 (02) :173-179

[7]

FRANKE W, 1955, Z PHYSIOL CHEM, V301, P90

[8] Detection of serum uric acid using the optical polymeric enzyme biochip system [J].

Huang, SH ;

Shih, YC ;

Wu, CY ;

Yuan, CJ ;

Yang, YS ;

Li, YK ;

Wu, TK .

BIOSENSORS & BIOELECTRONICS, 2004, 19 (12) :1627-1633

[9] Recurrent chemotherapy-induced tumor lysis syndrome (TLS) with renal failure in a patient with chronic lymphocytic leukemia - successful treatment and prevention of TLS with low-dose rasburicase [J].

Hummel, M ;

Buchheidt, D ;

Reiter, S ;

Bergmann, J ;

Adam, K ;

Hehlmann, R .

EUROPEAN JOURNAL OF HAEMATOLOGY, 2005, 75 (06) :518-521

[10]

IMSHENETSKII A A, 1971, Mikrobiologiya, V40, P269

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