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Modification of substrate specificity in single point mutants of Agrobacterium tumefaciens type II NADH dehydrogenase
Author(s) -
Desplats C.,
Beyly A.,
Cuiné S.,
Bernard L.,
Cournac L.,
Peltier G.
Publication year - 2007
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/j.febslet.2007.07.035
Subject(s) - agrobacterium tumefaciens , nadh dehydrogenase , biochemistry , mutant , dehydrogenase , enzyme , escherichia coli , nad+ kinase , substrate (aquarium) , wild type , chemistry , biology , protein subunit , transformation (genetics) , gene , ecology
Type II NADH dehydrogenases (NDH‐2) are monomeric flavoenzymes catalyzing electron transfer from NADH to quinones. While most NDH‐2 preferentially oxidize NADH, some of these enzymes have been reported to efficiently oxidize NADPH. With the aim to modify the NADPH vs NADH specificity of the relatively NADH specific Agrobacterium tumefaciens NDH‐2, two conserved residues (E and A) of the substrate binding domain were, respectively, mutated to Q and S. We show that when E was replaced by Q at position 203 the enzyme was able to oxidize NADPH as efficiently as NADH. Growth on a minimal medium of an Escherichia coli double mutant lacking both NDH‐1 and NDH‐2 was restored more efficiently when mutated proteins able to oxidize NADPH were expressed. The biotechnological interest of expressing such modified enzymes in photosynthetic organisms is discussed.