해녀콩(Canavalia lineata)잎에서 ornithine carbamoyltransferase(OCTase)를 부분정제 하여 canavanine대사와 연관된 일부성질과 효소활성의 세포내 분포를 조사하였다. 이 효소는 황산스트 토마이신 침전, ammonium sulfate 분획, DEAE-Sephacel 이온교환 크로마토그래피, 그리고 Sephacryl S-300 겔 크로마토그래피를 거쳐 정제한 결과 purification fold는 81.7이고 회수율은 6%이었으며 분자량은 120 kD로 산출되었고 활성의 최적 pH는 인산완충액의 경우 7.8이었다. 한편, 이 효소는 ornithine과 carbamoyl phosohate로부터 citrulline을 합성할 뿐만 아니라 canaline과 carbamoyl phosphate를 기지로 이용하여 ureidohomoserine을 합성할 수 있었다. 그리고 20mM carbamoyl phosphate일 때 ornithine에 대한 $K_m$은 14.1 mM이며 canaline에 대한 $K_m$은 5.5 mM이었고, 20mM ornithine일 때 carbamoyl phosphate에 대한 $K_m$은 10.6 mM으로 나타났다. OCTase 활성의 세포내 분포를 분별 원심분리와 Percoll 밀도구배법으로 조사한 결과 효소활성은 시토졸 분획(140,000 g 상등액)에 전체효소활성 중 61%가 집중되었고, 엽록체에서는 활성의 일부가 검출되었으나 미토콘드리아에서는 검출되지 않았다.
Ornithine carbamoyltransferase(OCTase) was partially purified and examined subcellular distribution of its activity from leaves of Canavalia lineata to study the role of OCTase in relation to canavanine biosynthesis. The purification steps involved streptomycin sulfate precipitation, ammonium sulfate fractionation, DEAE-Sephacel column chromatography, and Sephacryl S-300 gel chromatography. OCTase was purified 81.7 fold with 6% recovery. Molecular weight of the enzyme was estimated to be 120 kD and the optimum pH was 7.8 in potassium phosphate buffer. The partially purified enzyme produced citrulline from ornithine an carbamoyl phosphate, and also catalyzed canaline and carbamoyl phosphate to ureidohomoserine. The value of $K_m$ for ornithine and canaline were calculated to be 14.1 mM and 5.5 mM respectively with 20 mM carbamoyl phosphate, and $K_m$ for carbamoyl phosphate was 10.6 mM with 20 mM ornithine. The cytosolic fraction (140,000 g supernatant) by differential centrifugation contained 61% of the total OCTase activities. The fraction contained chloroplasts showed a little enzymatic activity (0.8%) after Percoll density gradient centrifugation, while no enzymatic activity was detected at mitochondrial fraction.
