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- Title
Kinetic Analysis of the Activation-and-Inhibition Dual Effects of Cobalt Ion on Thermolysin Activity.
- Authors
Hashida, Yasuhiko; Inouye, Kuniyo
- Abstract
Thermolysin activity as well as its stability is remarkably enhanced by high concentration of neutral salts consisting of Na+, K+, Cl− and Br− in the synthesis and hydrolysis of N-carbobenzoxy-L-aspertyl-L-phenylalanine methyl ester and hydrolysis of N-[3-(2-furyl)acryloyl]-glycyl-L-leucine amide (FAGLA) [Inouye, K. (1992) J. Biochem. 112, 335–340]. However, effect of divalent salts on thermolysin activity has not been investigated systematically. In this study, effect of Co2+ ion on thermolysin activity in the hydrolysis of FAGLA was examined. Thermolysin activity increased 3–4 times with increasing the Co2+ concentration to 2 mM, but the enhanced activity was considerably reduced with higher Co2+ concentration (2–18 mM). The activation-and-inhibition dual effects of Co2+ ion were analysed kinetically. Release of the catalytic Zn2+ ion from thermolysin, concomitantly occurred with the Co2+-dependent activation, was measured with a Zn2+-specific fluorescent probe. This indicates that the activation is caused by substituting Co2+ ion for the catalytic Zn2+ ion. Meanwhile, the Co2+-dependent activation was inhibited competitively by Zn2+ ion (0.1–1.0 μM) added, similarly to that it is inhibited by higher concentration of Co2+ ion. These lines of evidence provide a strategy for regulating thermolysin activity with Co2+ and Zn2+ ions.
- Subjects
AMINO acids; LEUCINE; HYDROLYSIS; MOLECULAR probes; FLUORESCENCE spectroscopy
- Publication
Journal of Biochemistry, 2007, Vol 141, Issue 6, p843
- ISSN
0021-924X
- Publication type
Article
- DOI
10.1093/jb/mvm088