방선균으로부터 Hydantoinase의 생산

Production of Hydantoinase from Streptomyces sp.

Hydantoinase(EC3.5.2.2) 생산을 위해 토양에서 분리한 균주 Y -183을 배양시킨 결과 Glycerol이 탄소원으로 양호했으며, 질소원으로는 혼합된 형태의 질소원이 요구되었다. 특히 Yeast ext.와 Beef ext.가 효소생성에 크게 영향을 미쳤고, Hydantoin 유도체 및 Pyrimidine 유도체의 영향을 조사한 결과 Uracil이 가장 양호한 결과를 나타냈다. 배양시간은 방선균의 특성상 72시간의 배양이 필요했으며 효소의 생성은 24시간 이후에 왕성하였다. 생성된 Hydantoinase는 균체내에 효소를 축적했으며 이들은 추출하여 기질로서 PL-5-phenylhydantion 등과 반응시킨 결과 N-carbamyl-D-amino acids가 생산되는 것이 분석결과 확인되었다.

In order to investigate hydantoinase-producing strain of the genus Streptomyces, 523 strains of Streptomycee sp. isolated from soils were cultivated in various media and conversion activity of the enzyme was measured to DL-5-phenylhydantoin. A number of strains producing hydantoinase were detected and among them, the strain of Streptomyces sp. Y-183 was selected as a most powerful strain to producing the enzyme. The optimal culture conditions for the production of hydantoinase of the strain were studied, and it was found that almost all hydantoinase activity was produced in the cell fraction. The maximum activity of the enzyme, 17.8 unitstg of dried cells weight, was obtained when the strain was cultured at $30^{\circ}C$ for 72 hr in a medium containing 1.0% of glycerol. 0.5% of yeast extract. 0.5% of soytone, 0.5% of beef extract, 0.6% of KCI, 0.002% of $K_2HP0_4, 0.25% \;of \;CaC0_3, \;0.0002% \; of \; ZnSO_4, \; 0.0002%\; of\; FeS0_4$, and 0.4% of uracil as an inducer, and the pH of culture broth was adjusted ranging from 7.0 to 7.5.