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KMID : 0578319940040020149
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Molecules and Cells
1994 Volume.4 No. 2 p.149 ~ p.154
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Identification and Purifiation of a RecA-like Protein from Fission Yeast Schizosaccharomyces pombe
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Lee, Jung Sup
Chun, Min Suck/Ahn, Kyung Sik/Hahn, Jae Kyu/Jang, Yeun Kyu/Park, Jong Kun/Kim, Sung Jun
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Abstract
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We have previously reported that SOS-like functions and/or related systems may exist in Schizosaccharomyces pombe (Lee and Park, 1994). The present study has been performed for identification and purification of a RecA-like protein from S. pombe. A protein with 74 kDa in size was identified from S. pombe with polyclonal antibodies raised against the homogeneous RecA protein obtained from Escherichia coll. This RecA-like protein of S pombe was concentrated with ammonium sulfate and further purified with DEAE-cellulose and Affi-Gel 10 columns immobilized with anti-RecA IgG. When the cell extract was fractionated by DEAE-cellulose chromatography, most of the intact 74 kDa proteins were eluted in flow through, whereas three fragments presumably degraded from the intact proteins were bound to DEAE-cellulose resin and could be eluted with a linear gradient of NaCl. These results indicate that if the a RecA-like protein of S. pombe is degraded by some yeast proteases, the degraded proteins show different binding affinity to DEAE-cellulose resin. Immunoaffinity chromatography was finally employed to purify a RecA-like protein of S. pombe. From the purification with wet weight 80 grams of cell mass, approximately 150 f;g of RecA-like protein with more than 90% of homogeneity was obtained, as judged by silver staining.
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