Cloning, characterization and expression of the Zymononas mobilis eda gene that encodes 2-keto-3-deoxy-6-phosphogluconate aldolase of the Entner-Doudoroff pathway

Abstract

The eda gene that encodes 2-keto-3-deoxy-6-phosphogluconate aldolase of the Entner-Doudoroff pathway was cloned from Zymomonas mobilis by genetic complementation of an Escherichia coli mutant. The gene is present in a single copy on the Z. mobilis genome and is not tightly linked to the edd gene. Nucleotide sequence analysis of the eda region revealed that the structural gene is 627 bp long and capable of encoding a protein of 208 amino acids with a deduced molecular weight of 21,505. The eda gene is monocistronic and is transcribed from a single promoter. The transcriptional initiation site was determined and an improved consensus promoter sequence for Z. mobilis was derived. High-level expression of the eda gene can be attributed to very efficient translational initiation caused by the high quality of the ribosome-binding site and stability of the mRNA, which has a decay rate of 7.6 min. A comparison of highly expressed Z. mobilis genes indicated that the relative quality of the ribosome-binding sites of these genes might play an important role in determining the level of enzyme synthesis. This possibility is discussed with regard to the role of gene expression in co-ordinating the enzyme levels of the Entner-Doudoroff glycolytic pathway.