Center for Applied Biotechnology Studies, Department of Biological Science, California State University Fullerton, Fullerton, CA, USA.
Sci Rep. 2019 Oct 10;9(1):14589. doi: 10.1038/s41598-019-51126-8.
Vibrio anguillarum 531A, isolated from a diseased fish in the Atlantic Ocean, is a mixture composed of about 95 and 5% of highly pigmented cells (strain 531Ad) and cells with normal levels of pigmentation (strain 531Ac), respectively. Analysis of the V. anguillarum 531Ad DNA region encompassing genes involved in the tyrosine metabolism showed a 410-bp duplication within the hmgA gene that results in a frameshift and early termination of translation of the homogentisate 1,2-dioxygenase. We hypothesized that this mutation results in accumulation of homogentisate that is oxidized and polymerized to produce pyomelanin. Introduction in E. coli of recombinant clones carrying the V. anguillarum hppD (4-hydroxyphenylpyruvate-dioxygenase), and a mutated hmgA produced brown colored colonies. Complementation with a recombinant clone harboring hmgA restored the original color to the colonies confirming that in the absence of homogentisate 1,2-dioxygenase the intermediary in tyrosine catabolism homogentisate accumulates and undergoes nonenzymatic oxidation and polymerization resulting in high amounts of the brown pigment. Whole-genome sequence analysis showed that V. anguillarum 531 Ac and 531Ad differ in the hmgA gene mutation and 23 mutations, most of which locate to intergenic regions and insertion sequences.
从大西洋患病鱼类中分离出的鳗弧菌 531A 是由约 95%和 5%的高度色素细胞（菌株 531Ad）和色素正常水平的细胞（菌株 531Ac）组成的混合物。对鳗弧菌 531Ad DNA 区域中参与酪氨酸代谢的基因进行分析，发现 hmgA 基因内有 410bp 的重复，导致同源戊二烯 1,2-双加氧酶的翻译移码和提前终止。我们假设这种突变导致了 homogentisate 的积累，这种物质被氧化并聚合产生 pyomelanin。在大肠杆菌中引入携带鳗弧菌 hppD（4-羟基苯丙酮酸双加氧酶）和突变 hmgA 的重组克隆，会产生棕色的菌落。用携带 hmgA 的重组克隆进行互补，使菌落恢复到原来的颜色，这证实了在缺乏 homogentisate 1,2-双加氧酶的情况下，酪氨酸代谢的中间产物 homogentisate 积累，并发生非酶氧化和聚合，导致大量棕色色素的产生。全基因组序列分析表明，鳗弧菌 531Ac 和 531Ad 在 hmgA 基因突变和 23 个突变上存在差异，其中大多数突变位于基因间区域和插入序列。
