嗜水气单胞菌asd-1基因功能研究

Function Verification Asd-1 Gene of Aeromonas hydrophila

asd-1基因编码天冬氨酸半醛脱氢酶,该酶可影响革兰氏阴性菌的细胞壁正常合成。为了研究嗜水气单胞菌(Aeromonas hydrophila)asd-1基因的功能,采用Overlap PCR方法敲除该基因,结果显示asd-1基因的敲除明显降低了嗜水气单胞菌的生长速度,但是不会导致突变株对二氨基庚二酸(DAP)的强制需求,进一步对氨基酸序列分析表明asd-1蛋白质较迟缓爱德华氏菌、鲶爱德华氏菌的asd A蛋白质多出3个氨基酸,其中Asn-257导致asd A蛋白质中的关键活性位点Arg-267在asd-1蛋白质中改变为Val-267,该研究表明嗜水气单胞菌的asd-1基因突变显著降低其生长速度。

Aspartate β-semialdehyde deshydrogenase which participated in the cell wall synthesis was edcoded by asd-1 gene.In order to revealing the function of Aeromonas hydrophila asd-1 gene, this study deleted it by overlap PCR technology. The result showed that the bacteria growth rate decreased significantly when asd-1 gene deleted, but the obligate requirement for diaminopimelic acid（DAP） in asd-1 mutants was not changed. Amino acid sequencs alignment further showed that asd-1 protein had 370 amino acids, but the asd A protein of Edwardsiella tada and Edwardsiella ictalari just had 367 amino acids. The Asn-257 resulted the key site Arg-267 in asd A protein changed to Val-267 in asd-1 protein. It indicated that asd-1 gene mutant decreased the growth rate of Aeromonas hydrophila significantly.