Structure-Function Studies of Escherichia coli Biotin Synthase via a Chemical Modification and Site-Directed Mutagenesis Approach

2001 ◽  
Vol 130 (5) ◽  
pp. 627-635 ◽  
Author(s):  
L. Farh ◽  
S.-Y. Hwang ◽  
L. Steinrauf ◽  
H.-J. Chiang ◽  
D. Shiuan
Keyword(s):  
Escherichia Coli ◽  
Chemical Modification ◽  
Structure Function ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Biotin Synthase

scholarly journals Role of cysteine residues in ribonuclease H from Escherichia coli. Site-directed mutagenesis and chemical modification

1990 ◽  
Vol 271 (1) ◽  
pp. 59-66 ◽  
Author(s):  
S Kanaya ◽  
S Kimura ◽  
C Katsuda ◽  
M Ikehara
Keyword(s):  
Escherichia Coli ◽  
Chemical Modification ◽  
Steric Hindrance ◽  
Thiol Group ◽  
Site Directed Mutagenesis ◽  
Ribonuclease H ◽  
Cysteine Residues ◽  
Directed Mutagenesis ◽  
E Coli

The role of the three cysteine residues at positions 13, 63 and 133 in Escherichia coli RNAase H, an enzyme that is sensitive to N-ethylmaleimide [Berkower, Leis & Hurwitz (1973) J. Biol. Chem. 248, 5914-5921], was examined by using both site-directed mutagenesis and chemical modification. Novel aspects that were found are as follows. First, none of the cysteine residues is required for activity. Secondly, chemical modification of either Cys-13 or Cys-133 with thiol-blocking reagents inactivates the enzyme, but that of Cys-63 does not. Thus the sensitivity of E. coli RNAase H to N-ethylmaleimide arises not from blocking of the thiol group but from steric hindrance by the modifying group incorporated at either Cys-13 or Cys-133.

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