Quorum Sensing (QS) in Vibrio cholerae, ensures to co-ordinate its behavioural changes at the inter/intra species level to establish its infection. The LuxO, a well-known QS response regulator controls the global (three parallel) QS cascade of Vibrio cholerae. The structural/functional predictions of LuxO shows to have a receiver, central ATPase and a DNA binding domains and we have identified the ATP binding domain as our molecular target as its main function is to hydrolysis the ATP molecule (source of energy and phosphate group). In the present study, we have modelled ATP binding domain of LuxO based on the structure 3MOE (crystal structure of ATP-bound state of walker B mutant NtrC1 ATPase domain) as templates. High throughput virtual screening of 1 million compounds from the public database were done against our target, LuxO. Based on the Glide score, interaction sites and ADMET property, the top 10 best hits were identified. The compound, 3 [4-morpholine-anilio] carbonyl-2-pyrazine carboxylic acid (QSIVc) has a glide score of -11.873 with favourable interactions to the amino acids- G170, G172, K 173, R 357, and I 140 and Mg2+. The conserved residues, R 357 that lies in the arginine finger of the ATPase domain and G 170 and G172 in the Walker A motif were functionally characterized to aid in the hydrolysis of ATP. So the proposed structure-function relationship of QSIVc and their exact interaction pattern to inhibit the ATP hydrolysis would be of significant interest to develop an effective cholera therapy.