Effects of chemical modification on the enzyme activity and stability of bromoperoxidase from a Thai seaweed
2
Issued Date
2023
Copyright Date
1995
Language
eng
File Type
application/pdf
No. of Pages/File Size
xxix, 334 leaves : ill.(some col.)
Access Rights
restricted access
Rights Holder(s)
Mahidol University
Bibliographic Citation
Thesis (Ph.D. (Biochemistry))--Mahidol University, 1995
Suggested Citation
Jirasak Kongkiattikajorn Effects of chemical modification on the enzyme activity and stability of bromoperoxidase from a Thai seaweed. Thesis (Ph.D. (Biochemistry))--Mahidol University, 1995. Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/88404
Title
Effects of chemical modification on the enzyme activity and stability of bromoperoxidase from a Thai seaweed
Alternative Title(s)
ผลของการเปลี่ยนแปลงทางเคมีต่อกัมมันตภาพในการทำงานและเสถียรภาพของเอนไซม์โบรโมเปอร์ออกซิเดสจากสาหร่ายทะเลไทย
Author(s)
Advisor(s)
Abstract
Bromoperoxidase was extracted from Thai red seaweed, Polycarvernosa sp. and purified by using 0-60% ammonium sulphate precipitation, DE 52 column chromatography and fast protein liquid chromatography by using MonoQ column and Superosel2 column. Two isoenzymes, BPO1 and BPO2 were separated in the purification step of DE52 column chromatography. After purification by Superosel2 column the specific activity of BPO1 and BPO2 were 61.86 and 330.58 mU/mg, respectively. BPO1 and BPO2, were non-heme enzymes that could be activated 2.5 and 3.5 fold by V(,2)O(,5). Chemical modification of BPO1 by2,4,6-trinitrobenzenesulfonic acid and iodoacetamide caused an enhancement in the enzyme activity and stability while the modification by N-acetylimidazole, diethylpyrocarbonate, 5, 5-dithiobis (2-nitrobenzoic acid) and 1,2-cyclohexanedione did not cause any change in enzyme activity and stability. The modification of BPO1 by N-bromosuccinimide, 2-hydroxy-5nitrobenzyl bromide, carbodiimide and iodoacetic acid caused a decrease in the enzyme activity. Chemical modification of BPO2 by 2,4,6-trinitrobenzenesulfonic acid, N-bromosuccinimide, 2-hydroxy-5-nitrobenzyl bromide, diethylpyrocarbonate, carbodiimide, iodoacetic acid and iodoacetamide inactivated the enzyme activity while modification by N-acetylimidazole, o-nitrophenylsulfenyl chloride and 1,2-cyclohexanedione did not change the enzyme activity. None of the modifying agent used in this study increased the enzyme activity or stability. The modification of BPO1 by iodoacetamide enhanced the enzyme activity about 500% and enhanced thermostability in both acidic and alkaline condition. K(,m) values of iodoacetamide-modified BPO1 for monochlorodimedone, potassium bromide and hydrogen peroxide were 1.43 x 10(-5), 1.54 x 10(-2) and 9.09 x 10(-7) M respectively while the K(,m) values of the native enzyme were 2.94 x 10(-5), 2.17 x 10(-4) and 1.00 x 10(-4) M, respectively. BPO1 contained two isoenzymes, BPOl.l and BP01.2 were isolated by using non-denaturing electrophoresis. From amino acid analysis of iodoacetamide-modified BPOl.l and BP01.2, the modified lysyl residues were 14 and 16 residues of the total lysyl residues of 42 and 49.5 residues, respectively at 18 hours of incubation at 35 degree C pH 7.0 while the modified histidyl residues were 6 and 9.5 of the total histidyl residues of 30 and 21.6 residues, respectively. The results suggested that the modified lysyl and histidyl residues might be involved in the enhancement of the enzyme activity and stability of BPO1.
Degree Name
Doctor of Philosophy
Degree Level
Doctoral Degree
Degree Department
Faculty of Science
Degree Discipline
Biochemistry
Degree Grantor(s)
Mahidol University