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Novel affinity chromatography method for the efficient purification of recombinant Binder of SPerm homolog proteins
Author(s) -
Zarafshan Samin Sabouhi,
Manjunath Puttaswamy
Publication year - 2020
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.202000250
Subject(s) - sperm , escherichia coli , recombinant dna , capacitation , affinity chromatography , sephadex , chemistry , chromatography , ion chromatography , biochemistry , biology , genetics , gene , enzyme
In mammalian species, a family of proteins named the Binder of SPerm proteins, which are expressed in the male reproductive tract, have been shown to play a role in epididymal sperm maturation and sperm capacitation. Recently, one homolog from human and two homologs from mouse were characterized. In order to further investigate the biochemical activity of these proteins, efficient purification procedures are required to isolate the proteins. Since these proteins are produced in very minute quantities, we exploited the high capacity of Escherichia coli to produce larger quantities of recombinant proteins that were subsequently purified using affinity chromatography on a diethylaminoethyl‐Sephadex A‐25 column. Binder of SPerm proteins have been shown to interact with pseudo‐choline groups such as diethylaminoethyl through affinity rather than ionic interactions. The aim of the current study was to develop a novel method for purifying these recombinant proteins, produced in Escherichia coli cells. Diethylaminoethyl is positively charged and is a weak anion exchanger, but binder of sperm proteins interacts with affinity to this resin. This study presents a new, rapid, and cost‐effective purification method that provides with an exceptional purity level, which can be used to study their roles in mammalian fertilization.