Vallorani Luciana, Polidori Emanuela, Sacconi Cinzia, Agostini Deborah, Pierleoni Raffaella, Piccoli Giovanni, Zeppa Sabrina, Stocchi Vilberto
Istituto di Chimica Biologica 'Giorgio Fornaini', Università degli Studi di Urbino, via Saffi, 2-61029 Urbino (PU), Italy.
New Phytol. 2002 Jun;154(3):779-790. doi: 10.1046/j.1469-8137.2002.00409.x.
• NADP-glutamate dehydrogenase (NADP-GDH) from Tuber borchii was purified and the corresponding gene was cloned in order to elucidate the physiological role of the enzyme in this ectomycorrhizal fungus. • NADP-GDH was purified using an anion-exchange column followed by affinity chromatography. The complete gene was cloned from a 30-d-old-mycelium cDNA library and characterized. • T. borchii NADP-GDH appears to be physically and kinetically similar to those from other fungi and the deduced amino acid sequence of the gdh gene showed a significant similarity to other fungal NADP-dependent GDHs. Biochemical and Northern blotting analyses carried out with mycelia grown on different nitrogen sources clearly showed that the regulation of T. borchii NADP-GDH in response to different nitrogen sources was markedly different from the responses of the NADP-GDHs of other ascomycetes. Northern blotting analyses highlighted that the gdh gene was also expressed in the symbiotic phase. • The biochemical and molecular data suggest that the fungal NADP-GDH contributes to the primary nitrogen metabolism in the ectomycorrhizal tissues.
• 为阐明NADP - 谷氨酸脱氢酶（NADP - GDH）在这种外生菌根真菌中的生理作用，对来自博氏块菌的NADP - GDH进行了纯化，并克隆了相应基因。
• 使用阴离子交换柱随后进行亲和层析对NADP - GDH进行纯化。从30日龄菌丝体cDNA文库中克隆了完整基因并进行了表征。
• 博氏块菌NADP - GDH在物理和动力学上似乎与其他真菌的相似，gdh基因推导的氨基酸序列与其他真菌的NADP依赖性GDH有显著相似性。对在不同氮源上生长的菌丝体进行的生化和Northern印迹分析清楚地表明，博氏块菌NADP - GDH对不同氮源的响应调节与其他子囊菌的NADP - GDH的响应明显不同。Northern印迹分析突出显示gdh基因在共生阶段也有表达。
• 生化和分子数据表明，真菌NADP - GDH有助于外生菌根组织中的初级氮代谢。
