Transcription Regulation of aco Operon in Klebsiella pneumoniae

• Main Authors: Ling Yun Ko, 柯伶昀
• Other Authors: Hwan-You Chang
• Format: Others
• Language: en_US
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/29519586785735431080

碩士 === 國立清華大學 === 生命科學系 === 89 === AcoK is a transcription regulator of aco operon in Klebsiella pneumoniae. It belongs to large ATP binding regulators of the LuxR family and shows high homology with MalT. In order to understand how AcoK regulates its target genes, the promoter region of either acoABCD or acoK was fused with the reporter gene luxAB and its expression was measured. The activity of Paco was very low during logarithmic growth and it started to increase after entering early stationary phase. Acetoin was shown to function as an inducer for the expression of aco genes, whereas glucose exhibited a repression effect. The result is in contrast to the expression of acoK, which is not autoregulated. The catabolites of acetoin metabolism such as 2,3-butanediol and acetaldehyde as well as other carbohydrates including galactose, lactose, and glycerol did not affect the expression of Paco and PacoK, whereas maltose stimulates the expression of PacoK at 210 min. The activity of Paco appeared to be high at alkaline pH0. Regions required for the function of Paco and PacoK were also identified. Finally, site-directed mutagenesis and deletion analysis were performed to investigate the function of AcoK. The ATP-binding domain I was shown to be essential for the ability of AcoK to activate Paco. The Asp residue located at 416th amino acid was also found to be critical for Paco activity. In conclusion, we have identified several factors that affect Paco and PacoK, and domains in AcoK that are critical for its activity. The result provides important insight on the transcription mechanism of aco genes.