Heterogeneous inhibition of horseradish peroxidase activity by cadmium.
Inhibition of horseradish peroxidase (HRP) activity by cadmium was studied under steady-state kinetic conditions after preincubation of the enzyme with millimolar concentrations of Cd(2+) for various periods of time. The H(2)O(2)-mediated oxidation of o-dianisidine by HRP was used to assess the enzymatic activity. Cd(2+) was found to be either a noncompetitive inhibitor of HRP or a mixed inhibitor of HRP depending both on the duration of incubation with HRP and on Cd(2+) concentration. Furthermore, for the same inhibition type, K(i) values dropped as incubation time increased. These results suggested that Cd(2+) would slowly bind to the enzyme and progressively induce conformational changes. Spectrophotometric analysis showed that indeed Cd(2+) altered the heme Soret absorption band on binding HRP and exhibited a K(d) which decreased as the incubation time of HRP with Cd(2+) increased. Hill plots suggested a cooperative binding of up to three Cd(2+) ions per molecule of HRP. Thus, Cd(2+) binding to HRP resulted in progressive inhibition of enzymatic activity with a change in the inhibition type as the number of Cd(2+) ions per HRP molecule increased. Results also illustrated the potential danger of long-term exposure to heavy metals, even for enzymes with low affinity for them.
Laboratory for Life Sciences, Saadat Abade, Sarve Sharghi, 34, 19979 Tehran, Iran. keyhanie@ibb.ut.ac.ir
Jacqueline
J
Keyhani
Ezzatollah
E
Keyhani
Nahid
N
Einollahi
Dariush
D
Minai-Tehrani
Sekineh
S
Zarchipour
D001665:Binding Sites; D002104:Cadmium / Q000378:metabolism / Q000633:toxicity*; D002118:Calcium / Q000378:metabolism; D002384:Catalysis; D006735:Horseradish Peroxidase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics
D002104:Cadmium; D006735:Horseradish Peroxidase; D002118:Calcium