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Literature summary for 3.4.24.27 extracted from

  • Kusano, M.; Yasukawa, K.; Hashida, Y.; Inouye, K.
    Engineering of the pH-dependence of thermolysin activity as examined by site-directed mutagenesis of Asn112 located at the active site of thermolysin (2006), J. Biochem., 139, 1017-1023.
    View publication on PubMed

Application

Application Comment Organism
synthesis introduction of ionizing residues into the active site of enzyme as a means of modifying its pH-activity profile Bacillus thermoproteolyticus

Protein Variants

Protein Variants Comment Organism
N112A no enzymic activity in supernatant of cells expressing mutant Bacillus thermoproteolyticus
N112D supernatants of cells expressing mutant show 18% of wild-type activity Bacillus thermoproteolyticus
N112E supernatants of cells expressing mutant show 5% of wild-type activity Bacillus thermoproteolyticus
N112H no enzymic activity in supernatant of cells expressing mutant Bacillus thermoproteolyticus
N112K no enzymic activity in supernatant of cells expressing mutant Bacillus thermoproteolyticus
N112R no enzymic activity in supernatant of cells expressing mutant Bacillus thermoproteolyticus

Organism

Organism UniProt Comment Textmining
Bacillus thermoproteolyticus P00800
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-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
N-carbobenzoxy-L-Asp-L-Phe-methyl ester + H2O
-
Bacillus thermoproteolyticus N-carbobenzoxy-L-Asp + L-Phe-methyl ester
-
?
N-[3-(2-furyl)acryloyl]-Gly-L-Leu amide + H2O
-
Bacillus thermoproteolyticus ?
-
?

pH Range

pH Minimum pH Maximum Comment Organism
5.5 8.5 for hydrolysis of neutral substrate N-[3-(2-furyl)acryloyl]-glycyl-L-leucine amide, wild-type and mutants N112D, N112E exhibit bell-shaped pH-dependence. For hydrolysis of negatively charged substrate N-carbobenzoxy-L-Asp-L-Phe methyl ester, wild-type shows bell-shaped pH-dependence, for mutants N112D and N112E, pH-dependence of the ratio kcat/Km decreases with increase in pH from 5.5 to 8.5 Bacillus thermoproteolyticus