Liu Y, Murata H, Chatterjee A, Chatterjee A K
Department of Plant Pathology, University of Missouri, Columbia 65211.
Mol Plant Microbe Interact. 1993 May-Jun;6(3):299-308. doi: 10.1094/mpmi-6-299.
Erwinia carotovora subsp. carotovora strain Ecc71 produces an array of extracellular enzymes including pectate lyase (Pel), polygalacturonase, cellulase, and protease. In strain Ecc71, these enzymes are coregulated by aepA, which encodes an activator of extracellular protein production (H. Murata, J. L. McEvoy, A. Chatterjee, A. Collmer, and A. K. Chatterjee, Mol. Plant-Microbe Interact, 4:239-246, 1991). The nucleotide sequence of a 2.7-kb aepA+ DNA segment revealed an open reading frame (ORF) of 1,395 bp which matches with the size of the aepA transcript determined by Northern blot analysis. aepA is predicted to encode a protein of 465 amino acid residues with a molecular mass of approximately 51 kDa and a pI of 6.52. The occurrence of a putative signal sequence and several hydrophobic domains suggest membrane localization of AepA. An aepA-lacZ operon fusion was constitutively expressed in E. coli (DH5 alpha) but inducible by pectate and celery extract in E. c. subsp. carotovora (AC5006). These findings suggest that aepA expression may be negatively regulated in E. c. subsp. carotovora. By assaying for the transcript of pel-1, which species a major secreted Pel species in strain Ecc71, and by following the expression of a pel1-lacZ operon fusion we determined that AepA activates pel-1 transcription. The characteristics of aepA including the lack of homology with other prokaryotic regulatory genes indicate that aepA encodes a novel regulatory protein required for extracellular protein production. Whereas homologs of Ecc71 aepA occur in E. c. subsp. carotovora and E. c. subsp. atroseptica strains, activation of exoenzyme production is markedly stimulated by aepA in E. c. subsp. carotovora than in E. c. subsp. atroseptica.
胡萝卜软腐欧文氏菌胡萝卜软腐亚种Ecc71菌株能产生一系列胞外酶，包括果胶酸裂解酶（Pel）、多聚半乳糖醛酸酶、纤维素酶和蛋白酶。在Ecc71菌株中，这些酶由aepA共同调控，aepA编码一种胞外蛋白产生的激活因子（H. 村田、J. L. 麦克沃伊、A. 查特吉、A. 科尔默和A. K. 查特吉，《分子植物 - 微生物相互作用》，4:239 - 246，1991年）。一个2.7 kb的aepA⁺ DNA片段的核苷酸序列揭示了一个1395 bp的开放阅读框（ORF），其与通过Northern印迹分析确定的aepA转录本大小相符。预测aepA编码一个由465个氨基酸残基组成的蛋白质，分子量约为51 kDa，pI为6.52。推定信号序列和几个疏水结构域的存在表明AepA定位于细胞膜。一个aepA - lacZ操纵子融合体在大肠杆菌（DH5α）中组成型表达，但在胡萝卜软腐欧文氏菌胡萝卜软腐亚种（AC5006）中可被果胶酸和芹菜提取物诱导。这些发现表明aepA的表达在胡萝卜软腐欧文氏菌胡萝卜软腐亚种中可能受到负调控。通过检测pel - 1的转录本（pel - 1编码Ecc71菌株中一种主要分泌的Pel种类），并追踪pel1 - lacZ操纵子融合体的表达，我们确定AepA激活pel - 1转录。aepA的特征包括与其他原核调控基因缺乏同源性，这表明aepA编码一种胞外蛋白产生所需的新型调控蛋白。虽然Ecc71 aepA的同源物存在于胡萝卜软腐欧文氏菌胡萝卜软腐亚种和黑腐欧文氏菌菌株中，但与黑腐欧文氏菌相比，aepA在胡萝卜软腐欧文氏菌胡萝卜软腐亚种中对胞外酶产生的激活作用明显更强。
