Unkles S E, Zhou D, Siddiqi M Y, Kinghorn J R, Glass A D
School of Biology, University of St Andrews, St Andrews KY16 9TH, UK.
EMBO J. 2001 Nov 15;20(22):6246-55. doi: 10.1093/emboj/20.22.6246.
Aspergillus nidulans possesses two high-affinity nitrate transporters, encoded by the nrtA and the nrtB genes. Mutants expressing either gene grew normally on 1-10 mM nitrate as sole nitrogen source, whereas the double mutant failed to grow on nitrate concentrations up to 200 mM. These genes appear to be regulated coordinately in all growth conditions, growth stages and regulatory genetic backgrounds studied. Flux analysis of single gene mutants using 13NO3(-) revealed that K(m) values for the NrtA and NrtB transporters were approximately 100 and approximately 10 microM, respectively, while V(max) values, though variable according to age, were approximately 600 and approximately 100 nmol/mg dry weight/h, respectively, in young mycelia. This kinetic differentiation may provide the necessary physiological and ecological plasticity to acquire sufficient nitrate despite highly variable external concentrations. Our results suggest that genes involved in nitrate assimilation may be induced by extracellular sensing of ambient nitrate without obligatory entry into the cell.
构巢曲霉拥有两个高亲和力硝酸盐转运蛋白,由nrtA和nrtB基因编码。表达任一基因的突变体在以1-10 mM硝酸盐作为唯一氮源的培养基上能正常生长,而双突变体在高达200 mM的硝酸盐浓度下无法生长。在所研究的所有生长条件、生长阶段和调控遗传背景中,这些基因似乎受到协同调控。使用13NO3(-)对单基因突变体进行通量分析表明,NrtA和NrtB转运蛋白的K(m)值分别约为100和约10 microM,而V(max)值虽然因年龄而异,但在年轻菌丝体中分别约为600和约100 nmol/mg干重/小时。这种动力学差异可能提供必要的生理和生态可塑性,以便在外部浓度高度可变的情况下获取足够的硝酸盐。我们的结果表明,参与硝酸盐同化的基因可能通过对环境硝酸盐的细胞外感知而被诱导,而无需进入细胞。