Hiroyuki YOSHIKAWA

Study on purification methods of aspartate racemase from bivalves

Ryouhei YAMADA, Yoshio KERA, Shouji TAKAHASHI,


Shibata et al in this laboratory successfully purified aspartate racemase from the foot muscle of the blood shell Scapharca broughtonii, by the procedure including Na2SO4 precipitation, Blue Sepharose column, Mono S column and Sephacryl S-100 column chromatography. In this study, I made attempts to improve the procedure for more effective purification of the enzyme and obtained the following results:
1. Introduction of AMP-Sepharose column chromatography in place of Mono S column chromatography described above enabled larger scale purification.
2. Na2SO4 precipitation conducted at 30�� in place of room temperature allowed
a more rapid operation, which enhanced the purification efficiency of this step.
3. Washing the Blue Sepharose column with 6 M urea and 30% isopropyl alcohol every time before use improved the efficiency of the chromatography.


I also made an attempt to purify aspartate racemase from another bivalve species Scapharca subcrenata for the first time. The enzyme could not be obtained as a homogeneous protein because of the small amount of specimen available, but it was purified with higher efficiency than the enzyme of the blood shell at several steps of the procedure.