Cloning, Sequencing, and Characterization of the Intracellular Invertase Gene from Zymomonas mobilis

Abstract

The structural gene for the intracellular invertase El of Zymomonas mobilis strain Z6C was cloned in a 2.25-kb DNA fragment on pUSHl 1, and expressed in Escherichia coli HB101. The enzyme produced by the E. coli carrying pUSHll was purified about 1,122 fold to homogeneity 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 β-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. © 1991, Japan Society for Bioscience, Biotechnology, and Agrochemistry. All rights reserved.