Mutations in V84I & A184V of bla TEM cluster plays a pivotal role in the dynamics of Ω-loop leading to genesis of IR-TEM.

The Enterobacteriaceae family exhibits resistance to antibiotics by producing β-Lactamase. Mutations in β-Lactamase, have led to the generation of inhibitor resistant variants known as IR-TEM. In the present study, IR-TEM β-Lactamase of Enterobacter hormaechei and Enterobacter asburiae was compared. To investigate the mechanism behind the conferred mutation, we studied its interaction with Clavulanic acid, (β-Lactamase inhibitor) with different lineages of TEM and IR-TEM. We found that Clavulanic acid quickly left the binding pockets of both variants using molecular dynamics (MD) simulations. Interestingly, mutations at the V84I and A184V positions were found to drastically influence the protein dynamics. Mutating the residues at V84I and A184V positions by computational mutagenesis in Enterobacter hormaechei , it was observed that the residues on the Ω-loop as well as a few downstream residues were primarily involved in generating resistance towards inhibitors by conferring increased flexibility to the loop. This further strongly supports the notion that inhibitor resistance in β-Lactamase is conferred through allosteric regulation, wherein mutations in positions 84 and 184 may play an important role in regulating the movement of the Ω-loop. These two positions determine the lineage pattern between two clusters in TEM-1 and TEM-116. To date, no reports have been made regarding the importance of these mutations and their dynamics in Ω-loop.Communicated by Ramaswamy H. Sarma.