Cloning, sequencing, and characterization of the intracellular invertase gene from Zymomonas mobilis.

The structural gene for the intracellular invertase E1 of Zymomonas mobilis strain Z6C was cloned in a 2.25-kb DNA fragment on pUSH11, and expressed in Escherichia coli HB101. The enzyme produced by the E. coli carrying pUSH11 was purified about 1,122 fold to homogenicity with a yield of 4%. The molecular weight and substrate specificity of the enzyme were identical with those of the intracellular invertase E1 from Z. mobilis. The nucleotides of the cloned DNA were sequenced; they included an open reading frame of 1,536 bp, coding for a protein with a molecular weight of 58,728. The N-terminal amino acid sequence predicted was identical with the sequence of the first 20 N-terminal amino acid residues of the protein obtained by Edman degradation. Comparison of the predicted amino acid sequence of E1 protein with those of the four other known beta-D-fructofuranosidases from Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae indicated a stronger homology in the N-terminal portion than in the C-terminal portion.