Structural And Functional Studies Of Vaccine Candidate Protein UgpB From Mycobacterium Tuberculosis

Tuberculosis is still one of the most devastating human diseases,and with the incidence of TB/HIV co-infection and drug-resistentTB,the global TB care and control is facing a huge challenge.At present, the only vaccinefor the prevention of TB is the BCG vaccinewhich is just largely efficacious among children. The current regimen for TB treatment comprises a cocktail of3-4drugs taken daily over a period of at least six months, yetpoor adherence to treatment poses a great impediment to effective therapy. Meanwhile, the number of drug-resistentcases is constantly increasing, and even cases of TDR have been reported. Thus, various new anti-TB drugs and vaccines are urgently needed for controllingglobal TB ravage. Structural insights into the drug-target and vaccine candidate proteins of M. tuberculosiswill not only promote discovery of new anti-TB lead compounds and drugs, but also help illustrate the biochemical mechanisms underlying the pharmacology and biology of tuberculosis combat.ABC transporters constitute a ubiquitous superfamily of integral membrane proteins, most of which utilize the energy of ATP hydrolysis to translocate solutes across membranes, and can be categorized as importers and exporters according to the translocation direction.The prokaryotic importers mediate the transmembrane uptake of nutrients for bacterial growth and survival.The essential nutrients forM. tuberculosis inside human host and the identity of corresponding transporters remain unknown till now.UgpABCE is one of the five identified putative permeases for carbohydrate uptake, and is genetically predicated to be a G3P importer. Its periplasmic binding subunit, UgpB, was reported to be essential for the optimal growth of M. tuberculosis in vitro, and also a potential anti-TB vaccine candidate.Here we determined the high resolution structure of UgpB and performed ITC assay to test potential substrates of UgpB accordingly. Interestingly, M. tuberculosis UgpBwas shown to accept GPC, rather than G3P, as its substrate, indicating GPC as a possible main carbon and phosphate source for the growth and survival of M. tuberculosis inside the human phagosome. Structural comparison has revealed that Trp169in E.coli UgpB, which has been shown to be essential for G3P binding, is replaced by Leu205in M. tuberculosis UgpB.In addition, some other residues involved in G3P binding in E.coliUgpB are not conserved in M. tuberculosis UgpB either. Therefore, M. tuberculosis lost the binding affinity for G3P.Mutational analysis of M. tuberculosis UgpB Leu205revealed its essential role in GPC binding.Our ITC experiments also showed that UgpB is unable to bind G2P or maltose as a substrate. The work presented here will not only contribute to our understanding of the carbon and phosphate sources utilized by M. tuberculosis inside the human host, provide structural insights into the development of anti-TB drug and vaccine, butalso promote improvements in TB therapy.