Characterization of Anthranilate Synthetase from a 5-methyltryptophan Resistant Mutant（MR1） in Maize

옥수수 5-methyltryptophan 저항성 돌연변이주（MR1）의 Anthranilate Synthetase 특성

5-methyltryptophan（5MT） resistant mutant plants （MRl） were analyzed for characterization of anthranilate synthetase （AS） and tryptophan synthetase （TS） enzymes. The enzyme was measured in crude extracts from MR1 and control seedlings of Danggin inbred line. There was no significant difference in the level of AS between MR1 and control seedlings when grown on MS medium without 5MT. However, MR1 seedlings grown on MS medium with 25mg/L 5MT showed the level of AS twice higher than that of control seedlings. The activity of AS was inhibited to 50％ in untreated plants when 4mg /L L-tryptophan was added to their extracts. Extracts from MR1 plants required about four times higher concentration of amino acid to cause equal inhibition. In the TS assay, the activity observed in MR1 seedlings was four times higher than that of control seedlings. We have also isolated and sequenced the gene which encoding the tryptophan synthetase B subunit （TSB） from maize. The gene encodes polypeptides with high homology to TSB isolated from other plants, and is expressed in all the developmental stages examined. Northern hybridization analysis indicated that the gene expression in MR1 seedlings grown on MS medium showed a higher level than in control seedlings.

옥수수의 5-methyltryptophan 저 항성 돌연변이 주（MR1）로부터 anthranilate synthetase（AS）와 tryptophan synthetase（75）의 특성을 분석하였다. 대조구 식물로써 사용한 옥수수 품종 당진의 순계와 저항성주에 있어서 AS와 75의 효소 활성은 5-MT를 포함하지 않은 MS기본배지에서 생장시켰을 때는 차이를 보이지 않았다. 그러나, 25mg/L의 5-MT를 포함한 MS배지에서 생장한 MR1에 있어서 AS의 활성은 대조구보다 2배 높았다. 또한, 4mg/L의 L-tryptophan을 처리 했을때의 AS의 활성은 50％ 저해 되었다. MR1의 조추출물로부터 대조구와 동일한 활성저해율을 나타내기 위해서는 약 4배의 아미노산이 필요하였다. MR1의 75활성은 대조 식물보다 4배가 더 높았다. 그리 하여 tryptophan synthetase B subunit （TSB）를 encoding하는 유전자를 cloning하여 염기배열을 결정 하였다. TSB유전자는 상이한 기관으로부터 cloning된 TSB와 높은 상동성을 보였으며, 모든 발육단계에서 발현하였다. 띠orthern hybridization분석에서 MR1의 TSB발현량은 대조식물보다 높게 나타났다