Active Lactamase Beta (LACTb)

Serine beta-lactamase-like protein (LACTB),an active-site serine protease,is the only mammalian mitochondrial homolog evolved from penicillin-binding proteins and β-lactamases (PBP-βLs) in bacteria.It is localized in the mitochondrial intermembrane space, where it assembles into filaments to regulate mitochondrial membrane organization and the efficiency of essential metabolic processes. Besides,C Reactive Protein (CRP) has been identified as an interactor of LACTB, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human LACTB and recombinant dog CRP. Briefly, LACTB was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to CRP-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-LACTB pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µL stop solution to the wells and read at 450/630nm immediately. The binding activity of recombinant human LACTB and recombinant dog CRP was shown in Figure 1, the EC50 for this effect is 0.152ug/mL.