Pectobacterium carotovorum subsp. carotovorum belongs to the Enterobacteriaceae family, which causes soft-rot disease in numerous plants worldwide resulting in significant economic losses. Results from our previous studies showed that the strain H-rif-8-6 produces low-molecular-weight bacteriocin (LMWB) Carocin S1. Interestingly, TH22–10, the caroS1K:Tn5 insertional mutant in H-rif-8-6, loses Carocin S1 producing ability, but still produces other LMWBs which the indicator strain SP33 can detect. The SP33 is one of the many strains that are sensitive toward the cytotoxic effects of Carocin S3K, but not Carocin S1. The result revealed that H-rif-8-6 is a multiple-bacteriocin producing strain. In this study, a 4.1-kb DNA fragment was isolated from the chromosomal DNA of Pcc strain, H-rif-8-6, by a DNA probe using the caroS1K gene as the template. DNA sequencing and analysis by GenBank revealed two complete open reading frames (ORFs), designated ORF1 and ORF2, which were identified within the sequence fragment. ORF1 and ORF2, similar to the identified carocin S2 genes, encode the killer (Carocin S3K) and the immunity (Carocin S3I) proteins, respectively, which were homologous to the colicin E3 gene. Carocin S3K and Carocin S3I were expressed, isolated, and purified in Escherichia coli BL21 after subcloning of the expression plasmid pGS3KI or pGSK3I. SDS-PAGE analysis showed that the relative masses of Carocin S3K and Carocin S3I were 95.6 kDa and 10.2 kDa, respectively. The results reveal that Carocin S3K has higher antimicrobial and specific antimicrobial activities for Pcc along with a nuclease activity than Carocin S3I. However, Carocin S3I inhibits the activity of Carocin S3K. Interestingly, a high concentration of Carocin S3I protein is also a DNA nuclease, and Carocin S3K also inhibits its activity. This study showed that another type of bacteriocin was found in Pectobacterium carotovorum. This new type of bacteriocin, Carocin S3, has the killer protein, Carocin S3K, and the immunity protein, Carocin S3I.

中文翻译：

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从多细菌素产生菌株 Pectobacterium carotovorum subsp. 中鉴定和表征细菌素 Carocin S3。胡萝卜素。

Pectobacterium carotovorum subsp。carotovorum 属于肠杆菌科，它在全世界许多植物中引起软腐病，造成重大经济损失。我们之前的研究结果表明，菌株 H-rif-8-6 产生低分子量细菌素 (LMWB) Carocin S1。有趣的是，H-rif-8-6 中的 caroS1K:Tn5 插入突变体 TH22-10 失去了 Carocin S1 产生能力，但仍产生指示菌株 SP33 可以检测到的其他 LMWB。SP33 是对 Carocin S3K 的细胞毒性作用敏感但对 Carocin S1 不敏感的众多菌株之一。结果表明，H-rif-8-6是一种多菌素产生菌株。在这项研究中，使用 caroS1K 基因作为模板，通过 DNA 探针从 Pcc 菌株 H-rif-8-6 的染色体 DNA 中分离出 4.1-kb DNA 片段。GenBank 的 DNA 测序和分析揭示了两个完整的开放阅读框 (ORF)，命名为 ORF1 和 ORF2，它们在序列片段中被鉴定出来。ORF1 和 ORF2 与已鉴定的 carocin S2 基因相似，分别编码与大肠杆菌素 E3 基因同源的杀伤蛋白（Carocin S3K）和免疫蛋白（Carocin S3I）。Carocin S3K 和 Carocin S3I 在表达质粒 pGS3KI 或 pGSK3I 亚克隆后在大肠杆菌 BL21 中表达、分离和纯化。SDS-PAGE分析表明Carocin S3K和Carocin S3I的相对质量分别为95.6 kDa和10.2 kDa。结果表明，与 Carocin S3I 相比，Carocin S3K 对 Pcc 具有更高的抗菌和特异性抗菌活性以及核酸酶活性。然而，Carocin S3I 抑制 Carocin S3K 的活性。有趣的是，高浓度的 Carocin S3I 蛋白也是一种 DNA 核酸酶，Carocin S3K 也会抑制其活性。这项研究表明，在食果肉杆菌中发现了另一种细菌素。这种新型细菌素 Carocin S3 具有杀伤蛋白 Carocin S3K 和免疫蛋白 Carocin S3I。