Potier S, Souciet J L, Lacroute F
Laboratoire de Génétique Physiologique, Institut de Biologie Moléculaire et Cellulaire du C.N.R.S., Strasbourg, France.
Mol Gen Genet. 1987 Sep;209(2):283-9. doi: 10.1007/BF00329655.
We replaced the URA2 gene by six different deleted alleles constructed in vitro by Bg/II digestion in order to correlate the genetic map with the restriction map and to define the regions coding for the different functions of the carbamylphosphate synthetase--aspartate transcarbamylase complex (CPSase-ATCase). We also enlarged the collection of ura2 point mutations by using a positive selection method based on resistance to the toxic accumulation of ureidosuccinic acid (USA). Of the new independent mutations nine mapped in the intermediary zone, a previously defined mutationless region localized between regions coding for CPSase and ATCase. This shows that the former definition resulted from analysis of a limited number of mutants (40). The study of an allele deleted in the intermediary zone shows that this sequence codes for a protein region necessary for the feedback inhibition of the CPSase-ATcase enzyme complex. The CPSase- ATCase- phenotype of 26 mutants resistant to USA accumulation shows the importance of the in vivo channelling of carbamylphosphate in the CPSase-ATCase complex for USA and subsequent pyrimidine biosynthesis. Finally, our results confirm that the CPSase and ATCase activities are separate functions.
我们用通过BglII消化体外构建的六个不同缺失等位基因取代URA2基因，以便将遗传图谱与限制图谱相关联，并确定编码氨甲酰磷酸合成酶-天冬氨酸转氨甲酰酶复合物（CPSase-ATCase）不同功能的区域。我们还通过基于对脲基琥珀酸（USA）毒性积累的抗性的正选择方法扩大了ura2点突变的收集。在新的独立突变中，有九个位于中间区域，这是一个先前定义的无突变区域，位于编码CPSase和ATCase的区域之间。这表明先前的定义是由于对有限数量的突变体（40个）进行分析得出的。对中间区域缺失的一个等位基因的研究表明，该序列编码CPSase-ATcase酶复合物反馈抑制所需的蛋白质区域。26个对USA积累有抗性的突变体的CPSase-ATCase表型表明，在CPSase-ATCase复合物中氨甲酰磷酸的体内通道化对USA和随后的嘧啶生物合成很重要。最后，我们的结果证实CPSase和ATCase活性是独立的功能。
