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Title: An easy two-step purification method for human leucocyte myeloperoxidase
Other Titles: İnsan lökosit miyeloperoksidazı saflaştırılması için kolay 2 basamaklı yöntem
Authors: Sarkarati, Bahram
Karaağaç Akyol, Tülay
Kılınç, Kamer
Keywords: Myeloperoxidase
Human leucocytes
Issue Date: Oct-2015
Publisher: Turkish Biochemistry Society
Source: Sarkarati, B., Akyol, T. K., & Kılınç, K. (2015). An easy two-step purification method for human leucocyte myeloperoxidase. [İnsan lökosit miyeloperoksidazı saflaştırılması için kolay 2 basamaklı yöntem] Turkish Journal of Biochemistry, 40(5), 410-416. doi:10.1515/tjb-2015-0035
Abstract: Objective: The object of this study is to describe a simple, rapid and cost effective method for purification of human leucocyte myeloperoxidase from a single donor. Myelopeoxidase (MPO) was purified by a two step procedure consisting of concanavalin-A Sepharose 4B affinity chromatography followed by CM-Sephadex cation exchange chromatography. Methods: Leucocytes from a single donor collected by leucopheresis were used in purification studies. MPO was solubilized and extracted from leucocytes by homogenization in phosphate buffer containing 1% HETAB (hexadecyltrimethylammonium bromide). MPO containing soluble material was applied onto concanavalin-A Sepharose 4B affinity gel, and was eluted with methyl-a-D-manno-piranoside. Fractions with MPO activity were pooled, dialyzed and applied onto CM-sephadex cation exchange gel, and was eluted from the column at weak cationic pH with linear NaCl gradient. Results: By the use of two chromatographic procedures, MPO was purified from human leucocytes with 70% yield. Purity of MPO was checked by determining the Reinheit Zahl (RZ) value (A(430)/A(280)). The RZ value of 0.86 indicated that the purified enzyme was highly homogenous as compared to reported experimental values (ranging from 0.82 to 0.88) and pure commercial enzyme with the RZ value of 0.84. Conclusion: In comparison with earlier purification methods, the purification method reported here has higher recovery rate and high purity together. Use of leucocytes with leucopheresis origin help us to omit the leucocyte isolation step and omitting of ammonium sulphate precipitation steps also help us to reduce the cost and is shortened the time of purification.
ISSN: 0250-4685
Appears in Collections:Scopus İndeksli Yayınlar Koleksiyonu / Scopus Indexed Publications Collection
Temel Tıp Bilimleri Bölümü / Department of Basic Medical Sciences
TR Dizin İndeksli Yayınlar / TR Dizin Indexed Publications Collection
WoS İndeksli Yayınlar Koleksiyonu / WoS Indexed Publications Collection

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