Maintenance and thermal stabilization of NADH dehydrogenase-2 conformation upon elimination of its C-terminal region

Development of an artificial enzyme with activity and structure comparable to that of natural enzymes is an important goal in biological chemistry. Respiratory NADH dehydrogenase-2 (NDH-2) of Escherichia coli is a peripheral membrane-bound flavoprotein, belonging to a group of enzymes with scarce structural information. By eliminating the C-terminal region of NDH-2, a water soluble version with significant enzymatic activity was previously obtained. Here, NDH-2 structural features were established, in comparison to those of the truncated version. Far-UV circular dichroism, Fourier transform infrared spectroscopy and limited proteolysis analysis showed that the overall structure of both proteins was similar at 30 degrees C. Experimental data agree with the predicted NDH-2 structure (PDB: 1OZK). The absence of C-terminal region stabilized in similar to 5-10 degrees C the truncated protein conformation. However, truncation impaired enzymatic activity at low temperatures, probably due to the weak interaction of the mutant protein with FAD cofactor. (C) 2012 Elsevier Masson SAS. All rights reserved.