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根据已发表的溶藻弧菌(Vibrio alginolyticus)血红素结合蛋白(Periplasmic Hemin-Binding Protein,HutB)基因序列设计引物,PCR扩增溶藻弧菌HY9901株的hutB基因,序列分析结果显示该基因全长870 bp,共编码289个氨基酸,分子量约为30.59 ku,PI为6.45。细胞定位、SignalP4.0、TMHMM Server 2.0和SoftB erry-Psite预测结果显示,hutB位于外周质中,存在信号肽切割位点,没有跨膜结构域,氨基酸序列含有1个cA MP和cG MP的蛋白激酶磷酸化位点,4个蛋白激酶C磷酸化位点等多个活性位点。系统进化树结果显示,溶藻弧菌hutB与坎氏弧菌(Vibrio campbellii)和哈氏弧菌(Vibrio harveyi)聚为一簇。qR T-PCR技术初步探究hutB在不同铁源下的表达量,结果表明,溶藻弧菌hutB在含有FeC l3的富铁培养基中,表达量与对照组相比差异不显著;在含有血红素的条件下表达量上调;在同时含有2-2’二联吡啶和血红素时上调极显著(P<0.01);在含2-2’二联吡啶的铁限制环境下,表达量下调极显著(P<0.01)。
The hutB gene of Vibrio alginolyticus HY9901 was amplified by PCR according to the published sequence of Hib genes of Vibrio alginolyticus (Hib), and the sequence analysis showed that the gene It is 870 bp long encoding a total of 289 amino acids with a molecular weight of about 30.59 ku and a PI of 6.45. Cell localization, SignalP4.0, TMHMM Server 2.0, and SoftBerry-Psite predicted that hutB was located in the periplasm with a signal peptide cleavage site, no transmembrane domain, and a protein with 1 cA MP and cG MP amino acid sequence Kinase phosphorylation sites, four protein kinase C phosphorylation sites and other active sites. Phylogenetic tree analysis showed that Vibrio alginolyticus hutB clustered with Vibrio campbellii and Vibrio harveyi. qR T-PCR technology initially explored the expression of hutB under different iron sources, the results showed that the expression of Vibrio alginolyticus hutB in iron-rich medium containing FeCl3 was not significantly different from the control group; (P <0.01). In the iron-restricted environment containing 2-2 ’dipyridyl, the expression level was down-regulated Significant (P <0.01).