EXPRESSION OF HUMAN LIVER ARGINASE IN ESCHERICHIA-COLI - PURIFICATION AND PROPERTIES OF THE PRODUCT

被引:54
作者
IKEMOTO, M
TABATA, M
MIYAKE, T
KONO, T
MORI, M
TOTANI, M
MURACHI, T
机构
[1] KYOTO UNIV,COLL MED TECHNOL,KYOTO 606,JAPAN
[2] TOSOH CORP,BIOTECHNOL RES LAB,AYASE,KANAGAWA 252,JAPAN
[3] KYOTO UNIV,FAC MED,DEPT CLIN SCI & LAB MED,KYOTO 606,JAPAN
[4] KUMAMOTO UNIV,SCH MED,INST MED GENET,KUMAMOTO 862,JAPAN
关键词
D O I
10.1042/bj2700697
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Arginase is an enzyme that catalyses the hydrolysis of arginine to urea and ornithine. It is abundantly present in the liver of ureotelic animals (i.e. those whose excretion is characterized by the excretion of uric acid as the chief end-product of nitrogen metabolism), but its purification has hitherto not been simple, and the yield not high. Starting with a partially truncated cDNA for human liver arginase recently made available, we constructed an expression plasmid that had tandemly linked tac promotors placed upstream of a full-length cDNA. By selecting Escherichia coli strain KY1436 as the host micro-organism, we established an efficient system for the production of human liver arginase protein. Chromatographies on CM-Sephadex G-150, DEAE-cellulose and Sephadex G-150, followed by preparative agar-gel electrophoresis, yielded 10 mg of apparently homogeneous enzyme protein from 1 g (wet wt.) of E. coli cells. E. coli-expressed human liver arginase had chemical, immunological and most catalytic properties indistinguishable from those of purified human erythrocyte arginase. However, E. coli-expressed arginase was a monomer of Mr 35,000, whereas the purified erythrocyte arginase was trimer of Mr 105,000. They differed also in pH- and temperature-stabilities. Gel-filtration experiments with these two purified arginases under various conditions, as well as with unfractionated human liver and erythrocyte cytosol preparations, indicated that the native form of human arginase should be of Mr 35,000, and that the trimeric appearance of human erythrocyte arginase after purification was an artifact of the purification procedures. It was thus concluded that, in Nature, the liver and erythrocyte arginases are identical proteins.
引用
收藏
页码:697 / 703
页数:7
相关论文
共 31 条
[1]   MOLECULAR FORMS OF HUMAN-LIVER ARGINASE [J].
BASCUR, L ;
CABELLO, J ;
VELIZ, M ;
GONZALEZ, A .
BIOCHIMICA ET BIOPHYSICA ACTA, 1966, 128 (01) :149-&
[2]   ARGINASE DEFICIENCY IN A 12-YEAR-OLD BOY WITH MILD IMPAIRMENT OF INTELLECTUAL FUNCTION [J].
BERNAR, J ;
HANSON, RA ;
KERN, R ;
PHOENIX, B ;
SHAW, KNF ;
CEDERBAUM, SD .
JOURNAL OF PEDIATRICS, 1986, 108 (03) :432-435
[3]   PURIFICATION AND PROPERTIES OF ARGINASE FROM HUMAN LIVER AND ERYTHROCYTES [J].
BERUTER, J ;
COLOMBO, JP ;
BACHMANN, C .
BIOCHEMICAL JOURNAL, 1978, 175 (02) :449-454
[4]   PLASMID VECTORS FOR THE SELECTION OF PROMOTERS [J].
BROSIUS, J .
GENE, 1984, 27 (02) :151-160
[5]   PURIFICATION, AFFINITY TO ANTI-HUMAN ARGINASE IMMUNOGLOBULIN-SEPHAROSE 4B AND SUBUNIT MOLECULAR-WEIGHTS OF MAMMALIAN ARGINASES [J].
BRUSDEILINS, M ;
KUHNER, R ;
SCHUMACHER, K .
BIOCHIMICA ET BIOPHYSICA ACTA, 1985, 840 (01) :79-90
[6]  
BURGESS RR, 1969, J BIOL CHEM, V244, P6168
[7]   HYPERARGININEMIA [J].
CEDERBAUM, SD ;
SHAW, KNF ;
VALENTE, M .
JOURNAL OF PEDIATRICS, 1977, 90 (04) :569-573
[8]  
CHANEY AL, 1962, CLIN CHEM, V8, P130
[9]   MOLECULAR CHARACTERISTICS OF CHICKEN LIVER ARGINASE [J].
GRAZI, E ;
MAGRI, E .
BIOCHEMICAL JOURNAL, 1972, 126 (03) :667-+
[10]   LIVER ARGINASE .3. PROPERTIES OF HIGHLY PURIFIED ARGINASE [J].
GREENBERG, DM ;
BAGOT, AE ;
ROHOLT, OA .
ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS, 1956, 62 (02) :446-453