Studies on endothelial nitric oxide synthase enzyme and its associated proteins in metabolic disorder

Abstract

Nitric Oxide Synthase (NOS) is an enzyme that catalyses the newlineproduction of Nitric Oxide (NO) from L-Arginine and molecular oxygen. newlineNO functions as a very important signalling molecule mediating many newlinebiological processes. Its deficiency is found to be associated with metabolic newlineand cardiovascular diseases. The current study addresses the dynamics of newlineendothelial Nitric Oxide Synthase (eNOS) enzyme, one of the three isoforms newlineof the NOS enzyme. Even though the NOS enzyme is being extensively newlinestudied, knowledge of the structure of each isoform and its dynamics may newlineprovide additional insights on its functional regulation. newlineThe eNOS protein comprises of the oxygenase domain in which the newlineheme active site resides and a reductase domain (FAD-NADP sub-domain newlineand FMN sub-domain) which serves as an electron transfer unit. A linker newlineregion connects the oxygenase and reductase domains. The FMN sub-domain newlinehas an auto-inhibitory loop (AI) region which keeps the enzyme in an inactive newlinestate. Activation of the enzyme requires the binding of a calcium binding newlineprotein calmodulin to the linker region of eNOS. Binding of calmodulin newlinerelieves the repression of the AI loop and activates the enzyme. newlineThe three-dimensional structure of the reductase domain of the newlinehuman eNOS enzyme is not available. Homology modeling of the FMN newlinecontaining reductase sub-domain of eNOS was performed in-silico, using the newlinereductase domain of the other two isoforms as structural templates. Homology newlinemodels were generated with and without the calmodulin protein binding newlinelinker region (CBL) that connects the oxygenase and reductase domains. newlineAn analysis of the secondary structure was performed and the modeled newlinestructures were validated before being used for further studies. newlinePhosphorylation is an important regulation mechanism that acts as newlinea switch to activate or inactive the enzyme newline newline