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Toward a New Generation of Glucose Isomerases Through Genetic Engineering
Author(s) -
Sicard P. J.,
Leleu J.B.,
Tiraby G.
Publication year - 1990
Publication title -
starch ‐ stärke
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.62
H-Index - 82
eISSN - 1521-379X
pISSN - 0038-9056
DOI - 10.1002/star.19900420108
Subject(s) - tetramer , isomerase , protein engineering , enzyme , gene , biochemistry , dimer , glucose 6 phosphate isomerase , sequence (biology) , mutagenesis , homology (biology) , chemistry , strain (injury) , stereochemistry , genetics , biology , computational biology , mutant , organic chemistry , anatomy
Streptomyces violaceoniger glucose isomerase (GI) was originally developed in 1973 when expectations about the future of HFCS in Europe were great. Improvement of the original strain CBS 409—73 obliged to consider the possible utilizations of genetic engineering as the best possible way for obtaining hyperproduction of the enzyme. The structure gene of GI was cloned and transferred in an integrating plasmid PUT 220 derived from pIJ 702. The sequence of the gene was determined and compared to the aminoacid sequence of the purified glucose isomerase. Crystallization of the enzyme was achieved and its 3‐D structure established; S. volaceoniger GI is normally existing as a tetramer with 4 identical sub‐units of 388AA each. The monomer includes a (alpha‐beta) 8 barrel sub‐structure and a large hydrophobic loop involved in the formation of dimers. Simultaneously, structural analysis of S. olivochromogenes GI revealed a great similitude with S. violaceoniger GI, though the former could exist as a dimer. In view of this situation and considering that because of a high level of homology, both enzymes presented significant differences in their functional properties, hybrid molecules were constructed and site directed mutagenesis utilized to create new GI species with modified properties.