Biochemistry

Candidatus Liberibacter asiaticus is a fastidious prokaryotic α-proteobacterium that has not been cultured as yet. It is the causative agent of Citrus Huanglongbing (HLB) is a disease that infects citrus plants causing huge losses to the citrus industry. The plants employ a variety of defense mechanisms to combat the pathogen attack which mainly involves the formation of reactive oxygen species (ROS) mainly hydrogen peroxide, peroxynitrite and organic hydroperoxides. The microbe on the other hand has antioxidant proteins that counter the reactive oxygen species. Peroxiredoxin (Prx) which is known to play a crucial role in peroxide detoxification is a super family of one of such antioxidant proteins. In this work, we have determined the crystal structure of Bacterioferritin Comigratory Protein (BCP) that belongs to 1-Cys Prx enzyme having peroxidatic/sulfenic acid cysteine (C-47) but lacks the resolving cysteine. Furthermore, we are investigating structural aspects to gain insight into the binding of different peroxide substrates at the active site. Site directed mutagenesis has been done to introduce the non-conserved resolving cysteine to study its effect on peroxidase activity in both wild type and mutant form. Biochemically, both BCP 1-cys and BCP 2-cys have shown peroxidase activity. DNA protection activity against oxidative damage and cell-line based assays has implicated its protective role towards the hypoxic condition of cells. Hence, the protein becomes a lucrative target for ligands which might be potential antimicrobials.

AbstractHuanglongbing (HLB) is a destructive citrus disease. The leading cause of HLB is Candidatus Liberibacter asiaticus. Fatty acid biosynthesis is essential for bacterial viability and has been validated as a target for the discovery of novel antibacterial agents. Enoyl−acyl carrier protein reductase (also called ENR or FabI and a product of the fabI gene) is an enzyme required in a critical step of bacterial fatty acid biosynthesis and has attracted attention as a target of novel antimicrobial agents. We determined the crystal structures of FabI from Ca. L. asiaticus in its apoform as well as in complex with b‐nicotinamide adenine dinucleotide (NAD) at 1.7 and 2.7 Å resolution, respectively, to facilitate the design and screening of small molecule inhibitors of FabI. The monomeric ClFabI is highly similar to other known FabI structures as expected; however, unlike the typical tetramer, ClFabI exists as a hexamer in crystal, whereas as dimer in solution, on the other hand, the substrate binding loop which always disordered in apoform FabI structures is ordered in apo‐ClFabI. Interestingly, the structure of ClFabI undergoes remarkable conformational change in the substrate‐binding loop in the presence of NAD. We conclude that the signature sequence motif of FabI can be considered as Gly‐(Xaa)5‐Ser‐(Xaa)n‐Val‐Tyr‐(Xaa)6‐Lys‐(Xaa)n‐Thr instead of Tyr‐(Xaa)6‐Lys. We have further identified isoniazid as a competitive inhibitor with NADH.

Summary ‘ Candidatus Accumulibacter’ is a biotechnologically important bacterial group that can accumulate large amounts of intracellular polyphosphate, contributing to biological phosphorus removal in wastewater treatment. Since its first molecular identification more than a decade ago, this bacterial group has drawn significant research attention due to its high abundance in many biological phosphorus removal systems. In the past 6 years, our understanding of Accumulibacter microbiology and ecophysiology has advanced rapidly, largely owing to genomic information obtained through shotgun metagenomic sequencing efforts. In this review, we focus on the metabolism, physiology, fine‐scale population structure and ecological distribution of Accumulibacter, aiming to integrate the information learned so far and to present a more complete picture of the microbiology of this important bacterial group.