scholarly journals The use of limited proteolysis to probe interdomain and active site regions of beta-galactosidase (Escherichia coli).

1988 ◽  
Vol 263 (4) ◽  
pp. 1848-1854
Author(s):  
L A Edwards ◽  
M R Tian ◽  
R E Huber ◽  
A V Fowler
Keyword(s):  
Escherichia Coli ◽  
Active Site ◽  
Limited Proteolysis ◽  
Beta Galactosidase

Effects of amino acid substitutions at the active site in Escherichia coli beta-galactosidase.

Genetics ◽  
1988 ◽  
Vol 120 (3) ◽  
pp. 637-644
Author(s):  
C G Cupples ◽  
J H Miller
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Active Site ◽  
Site Directed Mutagenesis ◽  
Amino Acid Substitutions ◽  
Lacz Gene ◽  
Wild Type ◽  
Mutant Proteins ◽  
Beta Galactosidase

Abstract Forty-nine amino acid substitutions were made at four positions in the Escherichia coli enzyme beta-galactosidase; three of the four targeted amino acids are thought to be part of the active site. Many of the substitutions were made by converting the appropriate codon in lacZ to an amber codon, and using one of 12 suppressor strains to introduce the replacement amino acid. Glu-461 and Tyr-503 were replaced, independently, with 13 amino acids. All 26 of the strains containing mutant enzymes are Lac-. Enzyme activity is reduced to less than 10% of wild type by substitutions at Glu-461 and to less than 1% of wild type by substitutions at Tyr-503. Many of the mutant enzymes have less than 0.1% wild-type activity. His-464 and Met-3 were replaced with 11 and 12 amino acids, respectively. Strains containing any one of these mutant proteins are Lac+. The results support previous evidence that Glu-461 and Tyr-503 are essential for catalysis, and suggest that His-464 is not part of the active site. Site-directed mutagenesis was facilitated by construction of an f1 bacteriophage containing the complete lacZ gene on a single EcoRI fragment.

scholarly journals Derived amino acid sequence and identification of active site residues of Escherichia coli beta-hydroxydecanoyl thioester dehydrase.

1988 ◽  
Vol 263 (10) ◽  
pp. 4641-4646 ◽  
Author(s):  
J E Cronan ◽  
W B Li ◽  
R Coleman ◽  
M Narasimhan ◽  
D de Mendoza ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Active Site ◽  
Active Site Residues
Sign in / Sign up

Export Citation Format

Share Document