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Cytotoxicity of apo bovine ?-lactalbumin complexed with La3+ on cancer cells supported by its high resolution crystal structure.

ABSTRACT: Cancer remains one of the biggest threats to human society. There are massive demands for compounds to selectively kill cancerous cells. Earlier studies have shown that bovine ? -lactalbumin made lethal to tumor cells (BAMLET) becomes cytotoxic against cancer cells in complex with oleic acid {Hoque, M. et. al., PLoSOne 8, e68390 (2013)}. In our study, we obtained bovine ?-lactalbumin complexed with lanthanum ion (La3+-B-?-LA) and determined its high resolution crystal structure. The natural calcium binding site of bovine ?-lactalbumin is replaced by lanthanum. The La3+ complex formation by B-?-apo-LA was also supported by various biophysical methods. Interestingly, our complex, La3+-B-?-LA exhibits much greater anticancer activity against breast cancer cells as compared to the reported BAMLET-oleic acid complex. This study shows that La3+-B-?-LA complex is preferentially more toxic to MCF-7 cells as compared to KB (oral cancer) and HeLa (cervical) cells, while almost non-toxic to the healthy cells that we studied. Our data indicates that the cytotoxicity of La3+-B-?-LA against cancer cells is through apoptotic path way. The higher anticancer activity of La3+-B-?-LA is attributable to the requisite structural changes induced in the protein by La3+ binding as supported by the crystal structure of the complex.

SUBMITTER: Yarramala DS

PROVIDER: S-EPMC6370903 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe enzyme dihydropteroate synthase (DHPS) participates in the de novo synthesis of folate cofactors by catalyzing the formation of 7,8-dihydropteroate from condensation of p-aminobenzoic acid with 6-hydroxymethyl-7,8-dihydropteroate pyrophosphate. DHPS is absent from humans, who acquire folates from diet, and has been validated as an antimicrobial therapeutic target by chemical and genetic means. The bacterium Burkholderia cenocepacia is an opportunistic pathogen and an infective agent of cystic fibrosis patients. The organism is highly resistant to antibiotics and there is a recognized need for the identification of new drugs against Burkholderia and related Gram-negative pathogens. Our characterization of the DHPS active site and interactions with the enzyme product are designed to underpin early stage drug discovery.ResultsAn efficient recombinant protein expression system for DHPS from B. cenocepacia (BcDHPS) was prepared, the dimeric enzyme purified in high yield and crystallized. The structure of the apo-enzyme and the complex with the product 7,8-dihydropteroate have been determined to 2.35 Å and 1.95 Å resolution respectively in distinct orthorhombic crystal forms. The latter represents the first crystal structure of the DHPS-pterin product complex, reveals key interactions involved in ligand binding, and reinforces data generated by other structural studies. Comparisons with orthologues identify plasticity near the substrate-binding pocket and in particular a range of loop conformations that contribute to the architecture of the DHPS active site. These structural data provide a foundation for hit discovery. An intriguing observation, an artifact of the analysis, that of a potential sulfenamide bond within the ligand complex structure is mentioned.ConclusionStructural similarities between BcDHPS and orthologues from other Gram-negative species are evident as expected on the basis of a high level of sequence identity. The presence of 7,8-dihydropteroate in the binding site provides details about ligand recognition by the enzyme and the different states of the enzyme allow us to visualize distinct conformational states of loops adjacent to the active site. Improved drugs to combat infections by Burkholderia sp. and related Gram-negative bacteria are sought and our study now provides templates to assist that process and allow us to discuss new ways of inhibiting DHPS.

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Cytotoxicity of apo bovine ?-lactalbumin complexed with La3+ on cancer cells supported by its high resolution crystal structure.

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