Enhancement of catalytic activity by gene truncation: Activation of L-aspartase from Escherichia coli

被引:7
作者
Jayasekera, MMK [1 ]
Saribas, AS [1 ]
Viola, RE [1 ]
机构
[1] UNIV AKRON,DEPT CHEM,AKRON,OH 44325
关键词
D O I
10.1006/bbrc.1997.7294
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
Aspartase front Escherichia coli is activated by proteolysis at the carboxy-terminal. A systematic study has been undertaken with the goal of identifying the amino acids in this region that influence the catalytic activity of aspartase. Stop codons have been introduced at various positions to prematurely truncate the aspA gene that encodes for aspartase by sequentially eliminating each. of the polar and charged amino acids in this region. The affinity of the enzyme for its substrate aspartic acid decreases systematically as each functionally significant amino acid is eliminated. However, enhanced catalytic activity (up to 2.5 times the k(cat) for native aspartase) is observed for those truncation mutants that end in a positively charged carboxyterminal amino acid. The precise position of the proteolytic activation of aspartase has been defined, and this covalent activation has been shown to be independent of the allosteric activation of aspartase that is also observed. (C) 1997 Academic Press.
引用
收藏
页码:411 / 414
页数:4
相关论文
共 24 条
[1]   L-ASPARTASE FROM ESCHERICHIA-COLI - SUBSTRATE-SPECIFICITY AND ROLE OF DIVALENT METAL-IONS [J].
FALZONE, CJ ;
KARSTEN, WE ;
CONLEY, JD ;
VIOLA, RE .
BIOCHEMISTRY, 1988, 27 (26) :9089-9093
[2]  
FISHER DB, 1973, J BIOL CHEM, V248, P4345
[3]  
GUEST JR, 1984, J GEN MICROBIOL, V130, P1271
[4]   STUDIES ON ASPARTASE [J].
ICHIHARA, K ;
KANAGAWA, H ;
UCHIDA, M .
JOURNAL OF BIOCHEMISTRY, 1955, 42 (04) :439-447
[5]  
IDA N, 1985, J BIOCHEM-TOKYO, V95, P35
[6]  
JAYASEKERA MM, 1997, BIOCHEMISTRY, V36
[7]  
JONES DH, 1993, PCR PROTOCOLS CURREN, P269
[8]   KINETIC-STUDIES OF L-ASPARTASE FROM ESCHERICHIA-COLI - SUBSTRATE ACTIVATION [J].
KARSTEN, WE ;
GATES, RB ;
VIOLA, RE .
BIOCHEMISTRY, 1986, 25 (06) :1299-1303
[9]   PURIFICATION OF ASPARTASE AND ASPARTOKINASE HOMOSERINE DEHYDROGENASE-I FROM ESCHERICHIA-COLI BY DYE LIGAND CHROMATOGRAPHY [J].
KARSTEN, WE ;
HUNSLEY, JR ;
VIOLA, RE .
ANALYTICAL BIOCHEMISTRY, 1985, 147 (02) :336-341
[10]  
KUCZENSKI R, 1973, J BIOL CHEM, V248, P2261