Wageningen University, Laboratory of Phytopathology, Droevendaalsesteeg 1, 6708 PB Wageningen, The Netherlands.
Fungal Genet Biol. 2011 Oct;48(10):990-7. doi: 10.1016/j.fgb.2011.06.002. Epub 2011 Jun 12.
D-galacturonic acid is the most abundant component of pectin, one of the major polysaccharide constituents of plant cell walls. Galacturonic acid potentially is an important carbon source for microorganisms living on (decaying) plant material. A catabolic pathway was proposed in filamentous fungi, comprising three enzymatic steps, involving D-galacturonate reductase, L-galactonate dehydratase, and 2-keto-3-deoxy-L-galactonate aldolase. We describe the functional, biochemical and genetic characterization of the entire D-galacturonate-specific catabolic pathway in the plant pathogenic fungus Botrytis cinerea. The B. cinerea genome contains two non-homologous galacturonate reductase genes (Bcgar1 and Bcgar2), a galactonate dehydratase gene (Bclgd1), and a 2-keto-3-deoxy-L-galactonate aldolase gene (Bclga1). Their expression levels were highly induced in cultures containing GalA, pectate, or pectin as the sole carbon source. The four proteins were expressed in Escherichia coli and their enzymatic activity was characterized. Targeted gene replacement of all four genes in B. cinerea, either separately or in combinations, yielded mutants that were affected in growth on D-galacturonic acid, pectate, or pectin as the sole carbon source. In Aspergillus nidulans and A. niger, the first catabolic conversion only involves the Bcgar2 ortholog, while in Hypocrea jecorina, it only involves the Bcgar1 ortholog. In B. cinerea, however, BcGAR1 and BcGAR2 jointly contribute to the first step of the catabolic pathway, albeit to different extent. The virulence of all B. cinerea mutants in the D-galacturonic acid catabolic pathway on tomato leaves, apple fruit and bell peppers was unaltered.
半乳糖醛酸是果胶的最丰富成分之一，果胶是植物细胞壁的主要多糖成分之一。半乳糖醛酸可能是生活在（腐烂）植物材料上的微生物的重要碳源。在丝状真菌中提出了一种分解代谢途径，该途径包含三个酶促步骤，涉及 D-半乳糖酸还原酶、L-半乳糖酸脱水酶和 2-酮-3-脱氧-L-半乳糖酸醛缩酶。我们描述了植物病原真菌 Botrytis cinerea 中整个 D-半乳糖醛酸特异性分解代谢途径的功能、生化和遗传特征。B. cinerea 基因组包含两个非同源半乳糖酸还原酶基因（Bcgar1 和 Bcgar2）、一个半乳糖酸脱水酶基因（Bclgd1）和一个 2-酮-3-脱氧-L-半乳糖酸醛缩酶基因（Bclga1）。它们的表达水平在含有 GalA、果胶或果胶作为唯一碳源的培养物中高度诱导。这四种蛋白质在大肠杆菌中表达，并对其酶活性进行了表征。B. cinerea 中的四个基因（分别或组合）的靶向基因替换，导致突变体在 D-半乳糖醛酸、果胶或果胶作为唯一碳源上的生长受到影响。在 Aspergillus nidulans 和 A. niger 中，第一次分解代谢转化仅涉及 Bcgar2 同源物，而在 Hypocrea jecorina 中，仅涉及 Bcgar1 同源物。然而，在 B. cinerea 中，BcGAR1 和 BcGAR2 共同参与分解代谢途径的第一步，尽管程度不同。所有 B. cinerea 突变体在番茄叶片、苹果果实和甜椒上 D-半乳糖醛酸分解代谢途径中的毒力均未改变。
