Analysis of predicted fungal proteomes revealed a large family of sequences that showed similarity to the Saccharomyces cerevisiae Class-I dihydroorotate dehydrogenase Ura1, which supports synthesis of pyrimidines under aerobic and anaerobic conditions. However, expression of codon-optimised representatives of this gene family, from the ascomycete Alternaria alternata and the basidiomycete Schizophyllum commune, only supported growth of an S. cerevisiae ura1Δ mutant when synthetic media were supplemented with dihydrouracil. A hypothesis that these genes encode NAD(P)⁺-dependent dihydrouracil dehydrogenases (EC 1.3.1.1 or 1.3.1.2) was rejected based on absence of complementation in anaerobic cultures. Uracil- and thymine-dependent oxygen consumption and hydrogen-peroxide production by cell extracts of S. cerevisiae strains expressing the A. alternata and S. commune genes showed that, instead, they encode active dihydrouracil oxidases (DHO, EC1.3.3.7). DHO catalyses the reaction dihydrouracil + O₂ → uracil + H₂O₂ and was only reported in the yeast Rhodotorula glutinis (Owaki in J Ferment Technol 64:205–210, 1986). No structural gene for DHO was previously identified. DHO-expressing strains were highly sensitive to 5-fluorodihydrouracil (5F-dhu) and plasmids bearing expression cassettes for DHO were readily lost during growth on 5F-dhu-containing media. These results show the potential applicability of fungal DHO genes as counter-selectable marker genes for genetic modification of S. cerevisiae and other organisms that lack a native DHO. Further research should explore the physiological significance of this enigmatic and apparently widespread fungal enzyme.

对预测的真菌蛋白质组的分析揭示了一大类序列，这些序列与酿酒酵母I 类二氢乳清酸脱氢酶 Ura1 具有相似性，该酶支持在需氧和厌氧条件下合成嘧啶。然而，当合成培养基补充有二氢尿嘧啶时，该基因家族的密码子优化代表（来自子囊菌链格孢菌和担子菌Schizophyllum commune）的表达仅支持酿酒酵母ura1Δ突变体的生长。假设这些基因编码 NAD(P) ⁺依赖的二氢尿嘧啶脱氢酶（EC 1.3.1.1 或 1.3.1.2）基于厌氧培养中缺乏互补而被拒绝。表达A. alternata和S. commune基因的S. cerevisiae菌株的细胞提取物的尿嘧啶和胸腺嘧啶依赖性耗氧量和过氧化氢表明，相反，它们编码活性二氢尿嘧啶氧化酶（DHO，EC1.3.3.7） . DHO 催化反应二氢尿嘧啶 + O ₂  → 尿嘧啶 + H ₂ O ₂并且仅在酵母红酵母中报道(Owaki in J Ferment Technol 64:205–210, 1986)。以前没有发现 DHO 的结构基因。表达 DHO 的菌株对 5-氟二氢尿嘧啶 (5F-dhu) 高度敏感，并且带有 DHO 表达盒的质粒在含有 5F-dhu 的培养基上生长过程中很容易丢失。这些结果表明真菌 DHO 基因作为对酿酒酵母和其他缺乏天然 DHO 的生物进行遗传修饰的反选择标记基因的潜在适用性。进一步的研究应该探索这种神秘且明显分布广泛的真菌酶的生理意义。