Physicochemical and kinetic characteristics of rhodanese from the liver of African catfish <Emphasis Type="Italic">Clarias gariepinus</Emphasis> Burchell in Asejire lake |
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Authors: | Omolara Titilayo Akinsiku Femi Kayode Agboola Adenike Kuku Adeyinka Afolayan |
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Institution: | (1) Department of Biochemistry, Obafemi Awolowo University, Ile-Ife, Nigeria; |
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Abstract: | Two forms of rhodanese were purified from the liver of Clarias gariepinus Burchell, designated catfish rhodanese I (cRHD I) and rhodanese II (cRHD II), by ion-exchange chromatography on a CM-Sepharose
CL-6B column and gel filtration through a Sephadex G-75 column. The apparent molecular weight obtained for cRHD I and cRHD
II was 34,500 ± 707 and 36,800 ± 283 Da, respectively. The subunit molecular weight determined by sodium dodecyl sulphate–polyacrylamide
gel electrophoresis was 33,200 ± 283 and 35,100 ± 141 Da for cRHD I and cRHD II, respectively. Atomic absorption spectrophotometric
analysis revealed that cRHD II contained a high level of iron (Fe), which presumably was responsible for the brownish colour
of the preparation. In contrast, no Fe was identified in cRHD I, and its preparation was colourless. Further characterization
of cRHD II gave true Michaelis–Menten constant (K
m) values of 25.40 ± 1.70 and 18.60 ± 1.68 mM for KCN and Na2S2O3, respectively, an optimum pH of 6.5 and an optimum temperature of 40°C. The Arrhenius plot of the effects of temperature
on the reaction rate consisted of two linear segments with a break occurring at 40°C. The apparent activation energy values
from these slopes were 7.3 and 72.9 kcal/mol. Inhibition studies on the cRHD II enzyme showed that the activity of the enzyme
was not affected by Mn2+, Co2+, Sn2+, Ni2+ and NH4
+, but Zn2+ inhibited the enzyme considerably. |
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Keywords: | |
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