TY - JOUR T1 - Substrate Utilization by Grape N-Carbamoylputrescine Amidohydrolase: The Final Enzyme in Putrescine Biosynthesis JF - American Journal of Enology and Viticulture JO - Am J Enol Vitic. SP - 137 LP - 144 DO - 10.5344/ajev.1997.48.2.137 VL - 48 IS - 2 AU - Mary Lou Mendum AU - Douglas O. Adams Y1 - 1997/01/01 UR - http://www.ajevonline.org/content/48/2/137.abstract N2 - N-Carbamoylputrescine (NCP), the final intermediate in putrescine synthesis from arginine in plants, is converted to putrescine by the enzyme NCP amidohydrolase. Published protocols for synthesizing NCP from cyanate and putrescine were found to yield NCP and a previously unsuspected side product which we identified as N-(4-aminobutyl)carbamic acid (NAC). NAC is formed from putrescine and carbonate arising from cyanate hydrolysis. Hydrochloric acid degrades NAC to putrescine but does not affect NCP. NCP was separated from putrescine by ion exchange, yielding a preparation of NCP without NAC. NCP without NAC, and radiopure NCP labeled with carbon-14 in the carbamoyl group were used to assay NCP amidohydrolase activity in crude extracts of grape callus tissue by trapping 14CO2 released by the extracts. NCP analogs with three to eight carbon side chains were prepared from the corresponding diamines with 14C in the carbamoyl moiety. Crude grape callus extracts released 14CO2 from each of the analogs, and unlabeled N- carbamoyldiaminohexane reduced the amount of 14CO2 evolved from carbamoyl-labeled NCP. Crude extracts also liberated 14CO2 from substituted ureas with three to six carbon side chains prepared from labeled cyanate and the corresponding amine. The unlabeled substituted urea derivatives were not clearly inhibitory to labeled NCP breakdown, suggesting that another enzyme activity in the crude extracts might be responsible for their degradation. ER -